Table of contents
Updated - April 28, 2026
The Huaier mushroom has been known for over 1,600 years and has been used successfully in cancer therapy since its cultivation.
history
It was first mentioned in a medical work around 240 AD Zhou Hou Fang of the doctor Ge Hong. The title refers to the period after the Zhou dynasty. It translates as „Manual of Emergency Medicine“, which focuses on prescriptions for first aid and is still referenced in medical contexts today.
It can also be found in the book Tang Ben Cao (Tang Dynasty), which was regarded as a reference work on herbal medicine.
The Huaier mushroom (Trametes robiniophila Murr) was used to treat chronic ailments, to speed up convalescence and for general strengthening. Among other things, it was said to promote blood circulation and eliminate the symptoms of the root causes of tumors.
Due to a lack of sufficient availability - the mushroom only grew in remote areas on the trunks of old specimens of the Chinese Sophora tree - it soon fell into oblivion.
Towards the end of the 1970s, Chinese scientists developed a method of cultivating the medicinal mushroom. Standardized production with consistent active ingredient quality has been possible since the early 1990s.
The Polysaccharide protein-complex (PS-T), consisting of polysaccharides and proteins, is the main active ingredient: a combination of a 6 Monosaccharides existing Heteropolysaccharide and one from 18 Amino acids composite protein.
Manufacturer
The manufacturer of Huaier granules is the company founded in 1995. Gaitianli Medicine Co, Ltd. based in Qidong, Jiangsu. The company maintains research, development and production facilities, test laboratories and warehouses on its approximately 130,000 square meter site with 1,700 employees. The production capacity amounts to 9,500 tons of Huaier mushroom and 250 million bags of granules.
The research focus is on the treatment of tumor and immune diseases. In studies, Huaier granules have shown promising results for treating cancer and blocking recurrences (source).
The Huaier mushroom is considered a research focus, especially in breast cancer therapy.
The product was initially produced as a reference substance for clinical studies (32% polysaccharides and 8% β-glucans) and was finally officially approved in China as an adjuvant therapeutic agent in oncology.
Today it is sold via pharmacies (PZN 19253502 - only 30% polysaccharides) and online platforms (Nutrimentas granules (identical concentration to the study content with 32% polysaccharides)) are distributed worldwide.
The Nutrimentas Granulate follows the scientific standard of the original manufacturer Gaitianli Medicine Co., Ltd. (32% polysaccharides) and contains 58% β-glucans!
There are supposedly „cheap“ providers of Huaier granules. The difference between the „expensive“ and „cheap“ product is that the expensive one is derived from the fruiting body of the Huaier mushroom, while the cheap one is produced from the mycelium on, for example, grain, which sometimes has only one-tenth the active ingredient content and contains 90% of undigested filler in solid-state fermentation.
In contrast, liquid fermentation, in which mycelium is cultivated in nutrient-rich liquid media, contains active ingredients that cannot be obtained from the fruiting body.
„Fruiting body“ refers to the mushroom as it is visually perceived, and „mycelium“ refers to the interior of the mushroom.
Studies
The most recent study from 2024, which has since been confirmed several times and was first published in 2022, is currently the Tanaka study by Dr. Manami Tanaka, M.D., Ph. D., Kanagawa, Japan, who works in biomedical research. His very detailed work, on the occasion of which the modRNA-vaccinations (Corona) ribosomal RNA (rRNA) in relation to the effect of the Huaier mushroom showed, contrary to the actual intention of his work, that cancer also responds to Huaier.
The study shows that taking Huaier extract in cancer patients has several effects: it normalizes ribosomal function, reduces the production of harmful spike proteins and prevents cancer recurrence with continued use.
The Huaier fungus has an „all-round“ effect - with the exception of brain tumors, as here the molecules probably cannot pass the blood-brain barrier due to the molecular size of the Huaier active substances with TP-1: 2300 kDa, HP-1: 30 kDa* - and is not selectively limited to just a few types of cancer.
This is because the active ingredients merely ensure the functional normalization of cell functions. This sounds very succinct, but is highly complex, as the study clearly demonstrates.
*kDa is used to indicate the mass of molecules, especially proteins. The unit Dalton (Da) is defined as the twelfth part of the mass of the carbon isotope 12C and is 1.66053906660(50) - 10-²⁷ kg. kDa is practically identical to kg/mol.
Further studies:
- Immunoregulatory effects of Huaier (Trametes robiniophila Murr) and relevant clinical applications.
- An extraction from Trametes robiniophila Murr. (Huaier) inhibits non-small cell lung cancer proliferation by targeting the epidermal growth factor receptor.
- Polysaccharides Produced by the Mushroom Trametes robiniophila Murr Boost the Sensitivity of Hepatoma Cells to Oxaliplatin via the miR-224-5p/ABCB1/P-gp Axis.
- Huaier Suppresses the Hepatocellular Carcinoma Cell Cycle by Regulating Minichromosome Maintenance Proteins
Function - explained for medical laypersons
Legal information: This information is for educational purposes and does not replace the advice of a specialist doctor/oncologist. Huaier granules are classified as a dietary supplement in Germany and not, as in China, as a medicinal product. Individual medical decisions should always be discussed with the treating oncologist.
Justified question from a reader: „... and why is the fungus not used by doctors and clinics?“
Because in Germany it is only approved as a dietary supplement (not, as in China, as a cancer drug), there is no billing option for it according to GOÄ or EBM (the fee schedules for doctors) and more money can be earned in the five-digit range with conventional chemotherapy drugs than with the comparatively „paltry“ annual costs of 2,200 euros for a Huaier treatment in the first year and only a few hundred euros in subsequent years.
Only a few doctors who operate outside of their purely conventional medical and billing-oriented colleagues are not only aware of the possibility of this therapy, but also support it.
Cancer cells undermine the body's control mechanism, the so-called. Hippo-Pathway (which determines whether a cell is healthy or defective and thus its cell death, the Apoptosis, ) and thus prevent apoptosis, which is why they continue to divide and multiply in an uncontrolled and uninhibited manner.
Huaier takes care of the repair of the Hippo-Pathways and thus enables the cell to resume its proper function, the switching on and off of various genes, and to correctly recognize and eliminate defective cells (Apoptosis).
To make matters worse, the so-called. Killer cells of the body (immune cells, NK cells) are exhausted in cancer patients and can no longer fulfill their task of destroying cancer cells.
Huaier activates these killer cells through his β-Glucans, This allows the immune system to be retrained and cancer cells such as metastases to be actively attacked and destroyed.
There are 8 main mechanisms by means of which the Huaier fungus successfully performs its amazing work, described below in excerpts and easy to understand.
1. the restoration of cell memory
A cell has specific tasks that are stored in the cell nucleus in the RNA (- RiboNuclein Acid = DNA - DeoxyriboNuclein Acid). A section of it codes for proteins that determine the function and structure of the cell by switching certain genes on or off.
To prevent this from happening, a gatekeeper (Hippo-Pathway) to ensure that the cell only fulfills its intended function. If it fails to do so, it is given around half an hour to correct the error. If it remains faulty, cell death (Apoptosis) is initiated to prevent a cell with incorrect information from multiplying.
However, if the gatekeeper fails, the incorrectly encoded cell will continue to divide inexorably.
The Huaier fungus reactivates the gatekeeper and thus restores control of cell function.
2. the genetic chaos
If the wrong genes are activated or deactivated or incorrectly switched on or off as a result of the incorrect information, other proteins are produced than required. The transcription factors are disrupted. As a result, the cell loses its assigned function.
However, genes are not simply switched on or off in binary form as in digital logic circuits, but are also finely adjusted, similar to a volume control, i.e. set to very quiet, quiet, medium, etc.. According to this setting, they ensure a reaction (expression) to the transmitted signals that is adapted to the respective situation.
The active ingredients of the Huaier mushroom reactivate thousands of genes correctly, bringing them back to their natural state, allowing the cell to resume its originally assigned function.
3. the metastasis brake
The signal paths PI3K, ACT and mTOR are used for intracellular communication, which determine growth, division behavior and metabolic processes, for example. If these are disrupted, the cell cannot develop its original function. If the signaling pathways are hyperactive, coordination of the processes is impossible, with the result that the cell goes out of control and divides rapidly (forms metastases).
Huaier inhibits this hyperactivity of the signaling pathways and thus prevents uncontrolled cell growth, including division and proliferation in the organism.
4. the miRNA control system
Compared to a car, the miRNA-control system (microRNA) represents the ABS control (spinning wheels are braked while gripping wheels are supplied with drive power). It prevents cells from skidding due to incorrect information.
Around 1,000 miRNAs are encoded in the human genome and control the switching state of genes.
For cancer Oncogenes (mutated genes that cause the growth of the cell to get out of control) are no longer slowed down, which promotes the growth and spread of the cancer.
At the same time Tumor suppressor genes (genes that control cell growth and division) are too strongly inhibited, which in turn allows the cancer to grow unchecked.
The Huaier fungus restores the defective genes to their intended switching position, which stops uncontrolled cell growth and prevents the defective cell from continuing to divide.
5. the exhausted immune system
The immune system is ultimately overwhelmed by the multiple malfunctions and can no longer adequately fight the cancer or keep it in check. The smallest infection, be it a cold, can develop into a full-blown pneumonia - with often fatal consequences.
The substances of the Huaier mushroom activate the NK cells (natural killer cells) and Macrophages (phagocytes), which kill cancer cells (Apoptosis) and absorb them in order to break them down. This allows the immune system to resume its protective function and fight the cancer effectively.
6 The wanderlust of cancer cells
Metastasis is the appearance of cancer cells in places other than the original tumor site. Normally, cancer cells adhere to the tissue where they originated. Over time, however, the EMT (Epithelial-mesenchymal transition) for the loss of the adhesive that holds the cell in place. As a result, it migrates around the organism, forming a movement protein that makes it even easier for it to move on.
The Huaier fungus inhibits this EMT process and thus prevents metastasis.
7 The supply system of cancer cells
Migrating cancer cells want to live and therefore form new blood vessels (Angiogenesis) to take care of themselves. This is how new cancerous tumors develop in various places in the body.
Huaier inhibits the growth factor VEGF, which is regulated by the hypoxia-inducible factor HIF-1α. This cuts off the supply to the resulting ulcers, which is why the tumor dies.
8. the defect in ribosomal RNA after chemotherapy
Chemotherapeutic agents damage the ribosomal RNA because they do not selectively target the DNA, but also affect other cellular structures.
Some chemotherapeutic agents, such as Actinomycin D, The DNA synthesis inhibitors, which are directly incorporated into the DNA double strand, not only prevent the formation of new DNA, but also the synthesis of RNA, including ribosomal RNA (rRNA), which is essential for protein production.
The cell thus loses its ability to synthesize proteins, which leads to cell death.
According to current research, the acute toxicity of Azacitidine mediated almost entirely via RNA damage. RNA damage apparently plays a central role in the effect of such drugs. Other substances, such as Anthracyclines, act by forming free radicals, which can damage both DNA and RNA.
This damage to the ribosomal RNA disrupts the function of the ribosomes, which are responsible for translation (Translation) of mRNA into proteins, which can ultimately lead to cell death.
The Huaier fungus repairs damage to the ribosomal structures, which helps healthy cells to regenerate, but causes cancer cells to die.
Application spectrum of Huaier
Huaier is applicable in the following types of cancer according to the aforementioned studies:
Breast cancer – Mammacarcinoma
- Quianquian Guo, e.a. – 01/05/2025 – „Effect of Huaier Granule on Prognosis of Breast Cancer: A Single-Center Propensity Score Matching Retrospective Study„
„214 patients with early-stage invasive breast cancer participated in this study, with 107 in the Huaier group and 107 in the control group. In the Kaplan–Meier analysis, the 2-year and 5-year DFS rates were significantly different between the Huaier group and the control group (Hazard Ratio [HR] 0.495; 95%confidence interval [CI] 0.257–0.953;
P = 0.023). The 2-year and 5-year OS rates also differed significantly (HR: 0.308; 95 % CI: 0.148–0.644;
P = 0.001). In the multivariate Cox regression, Huaier granules were associated with improved DFS (HR, 0.440; 95%CI, 0.223–0.868;
P = 0.018) and OS (HR, 0.236; 95%CI, 0.103–0.540;
P = 0,001).
In this retrospective study, Huaier granule improved the DFS and OS of patients with earlier invasive breast cancer.“ - Chen Li, a.o. – 09/18/2024 – „Huaier-induced suppression of cancer-associated fibroblasts confers immunotherapeutic sensitivity in triple-negative breast cancer„
„Remarkably, Huaier impairs the transition of CAFs into MyoCAFs by inhibiting SMAD2/3 phosphorylation, thereby overcoming MyoCAF-mediated T-cell dysfunction. This study underscores the considerable potential of Huaier as an effective adjuvant in cancer therapy.“ - Chen Li, a.k.a. – 06/16/2022 – „Huaier Induces Immunogenic Cell Death Via CircCLASP1/PKR/eIF2α Signaling Pathway in Triple Negative Breast Cancer„
„In the present study, we report that Huaier, an extract fromTrametes robiniophila Murr, inhibits TNBC progression by triggering ICD and leads to enhanced immunogenicity of cancer cells. Furthermore, the increased tumor-suppressive efficacy was largely dependent on CD8+ TIls dependent. The therapeutic effects of Huaier are closely related to ER stress above circCLASP1/PKR/eIF2α axis. Our results showed a promising role of traditional Chinese medicine in immune stimulation and provided supportive evidence for the application of Huaier as a potential ICD inducer in the treatment of TNBC patients.“
– 08/28/2020 – „Traditional Chinese biomedical preparation (Huaier Granule) for breast cancer: a PRISMA-compliant meta-analysis„
„In conclusion, the results of this meta-analysis suggest that the combination of Huaier granules and conventional treatment is effective in treating breast cancer patients. The clinical application of Huaier granules not only significantly enhanced the therapeutic effects of conventional treatment but also effectively improved the quality of life and immune function in patients with breast cancer.“ - Minghao Wang, e.a. – 02/12/2019 – „A Clinical Study on the Use of Huaier Granules in Post-Surgical Treatment of Triple-Negative Breast Cancer„
„Huaier granules could play an important role in the adjuvant postoperative therapy of TNBC patients, especially by effectively improving the DFS and OS of breast cancer patients in the middle to advanced stages.“ - Wei Wang, et al. – 11/08/2019 – „Huaier Suppresses Breast Cancer Progression Via linc00339/miR-4656/CSNK2B Signaling Pathway„
„Finally, we found that Huaier could inhibit the proliferation of breast cancer cells by modulating the linc00339/miR-4656/CSNK2B signaling pathway.“ - Zhang Y., e.a. – 05.2019 – „Efficacy of Huaier granule in patients with breast cancer„
„Our data showed that patients who received oral Huaier granules had a longer DFS (disease-free survival). Furthermore, Huaier granules might reduce serum tumor markers, improve the functional status, and the occurrence of emotional symptoms in breast cancer patients. Therefore, Huaier granules was an effective therapy for women with breast cancer.“ - JiJun Wang, b.a. – 11/17/2017 – „Huaier Extract Inhibits Breast Cancer Progression Through a LncRNA-H19/MiR-675-5p Pathway„
„In summary, the expression levels of H19, miR-675-5p, and CBL in breast cancer cell lines are affected by Huaier extract. Overexpression of H19 or miR-675-5p reverses the tumor-inhibiting effect of Huaier extract. However, eliminating H19 or miR-675-5p sensitizes breast cancer cells to Huaier extract. These data suggest that Huaier extract inhibits breast cancer cell proliferation and induces apoptosis via the lncRNA H19/miR-675-5p/CBL signaling pathway.“ - Yaming Li, e.a. – 02/01/2016 – „Huaier extract suppresses breast cancer by regulating tumor-associated macrophages„
„In summary, Huaier is an effective antitumor and immunomodulatory drug that could inhibit the infiltration of M2 macrophages into the breast cancer microenvironment, regulate TAM polarization, enhance phagocytosis, and reduce macrophage angiogenesis.“ - Ning Zhang, et al. – 07.16.2010 – „Huaier aqueous extract inhibits breast cancer cell proliferation by inducing apoptosis„
„In summary, our results indicated that Huaier extract can inhibit cell proliferation by inducing apoptosis and cell cycle arrest in breast cancer cells.“
Colon cancer – Colorectal carcinoma
- Qi Feng, e.a. – 01/17/2024 – „Huaier Regulates Oxaliplatin Resistance in Colorectal Cancer by Regulating Autophagy and Inhibiting the Wnt/β-catenin Signaling Pathway„
„This study has proven that Huaier can regulate autophagy, inhibit the Wnt/β-catenin signaling pathway, and reverse the drug resistance of OXA-resistant CRC cells.“ - Bo Li, e.a. – 08/03/2022 – „The treatment effects of Trametes Robiniophila Murr against colorectal cancer: A mini-review„
„In summary, Huaier extracts can inhibit the progression of CRC in various ways, including inducing apoptosis and inhibiting tumor cell proliferation, blocking epithelial-mesenchymal transition (EMT), attenuating CRC stem cell proliferation and differentiation, reducing tumor tissue vascular density, and strengthening the immune system. Therefore, Huaier could be an excellent candidate for increasing chemotherapy sensitivity while simultaneously enhancing the immune system and reducing side effects.“ - Yi-feng Zou, et al. – 08/26/2020 – „Effects of Huaier Extract on Ameliorating Colitis-Associated Colorectal Tumorigenesis in Mice„
„Huaier extract can reduce IBD-associated tumor development by suppressing pro-inflammatory cytokine levels and STAT3 stimulation.“ - Wen-Wen Sun, et al. – 07/11/2017 – „Killing effects of Huaier Granule combined with DC-CIK on nude mice transplanted with colon carcinoma cell line„
„Huaier Granule contains over 18 amino acids and a variety of minerals. It is now used to treat cancer by strengthening immunity and inhibiting tumor growth.“ - Tao Zhang, et al. – 01/21/2013 – „Huaier aqueous extract inhibits colorectal cancer stem cell growth partially via downregulation of the Wnt/β-catenin pathway„
„The present study showed that the aqueous Huaier extract is able to attack colorectal CSCs and inhibit spheroid formation potential as well as the ALDH-positive cell population. One of the mechanisms for the therapeutic effect of Huaier could be the downregulation of the Wnt/β-catenin self-renewal pathway. The present study suggests that the use of Huaier could be a good choice for treating CRC.“
LLiver cancer – Hepatocellular carcinoma (HCC)
- Zheng Jiaojiao, Wang Siying, Xia Lei, Sun Zhen, e.a. – 02/07/2025 – Hepatocellular Carcinoma: Signaling Pathways and Therapeutic Advances
„This report provides valuable insights into the treatment landscape of HCC and its potential application in various cancer types and pathological subtypes.“ - Junyu Wu, Guoyi Tang, Chien-Shan Cheng, et al. – 2024 – Traditional Chinese medicine for the treatment of hepatobiliary system cancers: from clinical evidence to drug discovery
„Among the TCMs currently under investigation, Huaier granule stands out as the most comprehensively studied, with three ongoing multicenter controlled trials evaluating its combination with TACE for HCC and with chemotherapy for PCC treatment, respectively (registered as NCT05660213, NCT06387368, and NCT06368063).“
Lung carcinoma – Bronchial carcinoma / Lung carcinoma
- Hooji Jin, October 11, 2023 – „Huaier suppresses cisplatin resistance in non-small cell lung cancer by inhibiting the JNK/JUN/IL-8 signaling pathway„
Huaier depresses Cisplatin Resistance and cancer stem cell properties in cisplatin-resistant NSCLC cells, both in vitro like in vivo. Mechanistically, Huaier could suppress the expression of interleukin-8 (IL-8) by inhibiting nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and activator protein-1 (AP-1), two key transcription factors responsible for IL-8 transcription activation.
Kaempferol was identified in Huaier as one of the small molecules that could suppress cisplatin resistance by inhibiting the phosphorylation and nuclear translocation of the proto-oncogene c-Jun (JUN) and by binding and inhibiting the kinase activity of c-Jun N-terminal protein kinase (JNK).“ - Xiangli Liu, et al. – 02.2021 – „Huaier demonstrates anti-cancer activities by inhibiting cell growth, migration, and energy metabolism in lung cancer through the PI3K/AKT/HIF-1α pathway.y“
„The in vivo data confirmed that Huaier evidently reduced tumor volume and tumor growth, and decreased glycolysis, glucose transport, and HIF-1α expression in tumor-bearing tissue. Our results suggest that Huaier exhibited anti-tumor effects both in vivo and in vitro, possibly through the PI3K/AKT/HIF-1α signaling pathway.“ - Ying-Ying Tian, etc. – „Effect of Huaier aqueous extract on growth and metastasis of human non-small cell lung cancer NCI-H1299 cells and its underlying mechanisms„
„The results showed that the aqueous Huaier extract inhibited the proliferation of NCI-H1299 cells and induced a cell cycle arrest at the phase. The aqueous S. Huaier extract promoted apoptosis of NCI-H1299 cells by down-regulating the expression of the anti-apoptotic protein Bcl-2. Furthermore, aqueous Huaier extract increased ROS levels and induced ferroptosis in NCI-H1299 cells. EMT played a crucial role in cancer metastasis. The aqueous Huaier extract reduced the migration ability of NCI-H1299 cells by inhibiting EMT of NCI-H1299 cells. In addition, this study found that aqueous Huaier extract inhibited the MAPK signaling pathway in human non-small cell lung cancer cells NCI-H1299, which might be one of Huaier's mechanisms in inhibiting the growth and metastasis of NCI-H1299 cells.“ - Yang Cheng, et al. – 03/22/2018 – „Huaier Granule extract inhibits the proliferation and metastasis of lung cancer cells by down-regulating the MTDH, JAK2/STAT3, and MAPK signaling pathways.„
„Huaier can inhibit the proliferation and metastasis of lung cancer cells by inhibiting the expression of MTDH and the activity of JAK2/STAT3 and MAPK signaling pathways. It has the potential for the treatment of lung cancer, given that Huaier possessed the characteristics of low toxicity and multi-target action, and significantly inhibited tumor proliferation and metastasis in vivo and in vitro.“ - Tangwei Wu, a.e.a. – 05/08/2014 – „Huaier suppresses proliferation and induces apoptosis in human pulmonary cancer cells via upregulation of miR-26b-5p„
„In conclusion, miR-26b-5pHuaier-mediated suppression of EZH2, β-catenin, and bcl-2 might be a central regulatory mechanism through which Huaier mediates reduced cell proliferation and increased apoptosis in lung cancer cells. Our study was the first to uncover the Huaier/
miR-26b-5pThe EZH2 signaling pathway in lung cancer cells, which offers new mechanisms for understanding the anti-tumor effect of Huaier and furthermore suggests a new basis for clinical treatment.“
Stomach cancer – Gastric adenocarcinoma / Stomach adenocarcinoma
- Hua-Xia Xie e.a. – 09.2015 – „Effect of Huaier on the proliferation and apoptosis of human gastric cancer cells„
„In summary, there are numerous signaling pathways that are important in Huaier-induced apoptosis. Among them, the PI3K/AKT signaling pathway is one of the most critical. The present results suggest that Huaier modulates the PI3K/AKT signaling pathway by inhibiting PI3K expression.“ - Jing Qi, a.e. – 07/08/2020 – „Huaier Granule combined with Tegafur Gimeracil Oteracil Potassium promotes stage IIb gastric cancer prognosis and induces gastric cancer cell apoptosis by regulating Livin.„
„Huaier granules in combination with Tegafur Gimeracil Oteracil Potassium could improve the prognosis of stage IIb gastric cancer. Huaier polysaccharides inhibited the proliferation of gastric cancer SGC-7901 cells and induced apoptosis of SGC-7901 cells by regulating Livin.“ - Zhiyuan Xu, et al. – 10/19/2017 – „Aqueous Huaier Extract Suppresses Gastric Cancer Metastasis and Epithelial-to-Mesenchymal Transition by Targeting Twist„
„Transwell and wound healing assays also confirmed that Huaier suppressed the migration and invasiveness of another GC cell line, MGC803.“ - Yiping Wang, et al. – 01/24/2019 – „Huaier n-butanol extract suppresses proliferation and metastasis of gastric cancer via the c-Myc-Bmi1 axis„
„In summary, we have shown that Huaier n-butanol extract inhibited GC cell proliferation, colony formation, migration, and invasion by down-regulating c-Myc and Bmi1.“ - Yunfu City, et al. – 06/15/2022 – „Huaier Inhibits Gastric Cancer Growth and Hepatic Metastasis by Reducing Syntenin Expression and STAT3 Phosphorylation„
„In summary, our study shows that HBE can inhibit the growth and metastasis of GC, especially liver metastasis, by inhibiting the syntenin/STAT3 signaling pathway and reversing EMT. This study provides a rationale for the use of HBE in the treatment of GC.“ - Daorui Hou, et al. – 08/21/2020 – „Efficacy and safety of Huaier granules combined with chemotherapy for gastric cancer: A protocol for systematic review and meta-analysis„
„In this study, we will conduct a systematic review and meta-analysis to provide further evidence on the efficacy and safety of Huaier granules in combination with chemotherapy.“
Nasopharyngeal carcinoma - Nasopharyngeal carcinoma
- RemarkNasopharyngeal carcinoma is mentioned in meta-analyses (e.g., in systematic reviews), but specialized clinical studies in connection with Huaier are currently not available.
Ovarian cancer - Ovarian carcinoma
- Xiaohui Yan, et al. – 05/08/2013 – „Huaier Aqueous Extract Inhibits Ovarian Cancer Cell Motility via the AKT/GSK3β/β-Catenin Pathway„
„In summary, Huaier not only inhibits cell growth by reducing proliferation and inducing apoptosis, but also reduces cell mobility in ovarian cancer cells via the AKT/GSK3β/β-Catenin signaling pathway. Given its low toxicity, Huaier is an attractive option for specifically targeting the invasion of epithelial ovarian cancer cells due to its effect on GSK3β/β-Catenin signaling.“
Metastases
Metastases are secondary tumors that form when malignant tumor cells break away from the primary tumor and migrate through the bloodstream or lymphatic system to other parts of the body, where they settle and multiply again. This applies to 0.01 to 0.1 % of circulating cancer cells.
This process significantly worsens the chances of curing cancer, as almost 90 percent of all cancer deaths are attributable to these metastases, not the primary tumor.
Preferred target regions for metastases are the liver, lungs, bones, brain, and lymph nodes.
Benign tumors do not metastasize.
Clinical effects of Huaier on metastases (especially liver)
A randomized multicenter clinical trial showed that Huaier granules reduced the recurrence rate in patients after curative resection of hepatocellular carcinoma (HCC).
Source: Chen Quian, et al. – BMJ Journals – Hepatology – Nov. 2018 – Effect of Huaier granule on recurrence after curative resection of HCC: a multicentre, randomised clinical trial
In gastric cancer xenografts, the combination of Trametes robiniophila Murr n-butanol extract with 5-fluorouracil showed a significant reduction in the risk of liver metastasis in vivo. PubMed.
Source: Jing-Li Xu, et al. – Frontiers in Pharmacology – 05/17/2022 – Trametes robiniophila Murr Sensitizes Gastric Cancer Cells to 5-Fluorouracil by Modulating Tumor Microenvironment
Molecular Mechanisms Against Metastasis
In vitro and in vivo experiments show that Huaier directly inhibits tumor proliferation, induces tumor cell death, prevents metastasis, and interferes with angiogenesis through various signaling pathways.
source: Jun Pan, et al. – Cancer Manag Res. – 02/14/2019 – Trametes robiniophila Murr: A Traditional Chinese Medicine with Potent Anti-tumor Effects
Huaier polysaccharide dose-dependently inhibited the proliferation, adhesion, migration, and invasion of HCC cells by reducing AEG-1 and N-cadherin expression, as well as enhancing E-cadherin.
source: Jiasheng Zheng, et al. – International Journal of Biological Macromolecules – March 2014 – Huaier polysaccharides suppress hepatocarcinoma MHCC97-H cell metastasis via inactivation of the EMT and AEG-1 pathways.y
Specific lung metastasis results
Huaier polysaccharide (TP-1) significantly inhibited tumor growth and lung metastasis in mice with HCC tumors without toxicity, while improving the proliferation of immune organs (spleen, thymus).
source: Cong Li, et al. – International Journal of Biological Macromolecules – 04.2015 – Huaier polysaccharide restrains hepatocellular carcinoma growth and metastasis by suppressing angiogenesis
Recent data (2024-2025)
A prospective cohort study showed that Huaier granules significantly prolong progression-free survival and reduce the risk of metastasis by 47%.
source: Hui Li, et al. – Frontiers in Pharmacology – 03/27/2025 – Enhancing survival outcomes in unresectable hepatocellular carcinoma: A prospective cohort study on the effects of Huaier granules with targeted therapy plus immunotherapy
After microwave ablation, long-term application of Huaier granules showed improved 3- and 5-year survival rates and reduced risk of extrahepatic metastasis.
source: Kailing Xi, et al. – Frontiers in Pharmacology – 05/07/2024 – Long-term oral administration of Huaier granules improves survival outcomes in hepatocellular carcinoma patients within Milan criteria following microwave ablation: a propensity score matching and stabilized inverse probability weighting analysis
Current scientific evidence focuses on HCC, while data on metastases from other organs to the liver are limited.
Active ingredients
The main active ingredients of the Huaier mushroom are divided into
1. β-glucans (beta-glucans) - 20-30% of the extract
- Polysaccharides with 1,3- and 1,6-glycosidic bonds
- Activate Toll-like Receptors (TLR2, TLR3, TLR6) on immune cells
- Stimulate Natural Killer Cells (NK cells) and Macrophages
- Increase TH1 cytokine production (IFN-γ, IL-2, TNF-α)
2. polysaccharides (30-40% of the extract in total)
- Modify the intestinal microbiota
- Promote the production of short-chain fatty acids (SCFAs)
- This activates G protein-coupled receptors (GPR43, GPR109A)
- Leads to epigenetic changes in immune cells
3. bioactive metabolites
- Polysaccharides with branched structure
- Triterpenes
- Phenolic compounds with antioxidant effect
When does taking Huaier granules have an effect?
The intake should be in direct connection with
- conventional surgery (accelerates wound healing)
- Chemotherapy (regenerates ribosomal RNA, prevents side effects)
- Irradiation
(after prior discussion with the treating oncologist and his knowledge of these contents) - hormone therapy, as there are no known interactions
- immunotherapy, due to synergistic effect
If the Huaier granules are taken regularly at the recommended dosage, the following effects can be observed:
Day 1-7:
- β-Glucans activate macrophages & NK cells
- First immune response is launched
Week 1-2:
- Transcription factors are reactivated
- First gene expression changes in cancer cells
Week 2-4:
- Massive gene expression conversion (1000s of genes)
- Hippo-Pathway is being repaired
- First apoptosis (cell death) in cancer cells
Week 4-12:
- EMT is blocked (metastasis prevention)
- Angiogenesis is inhibited (tumor starves)
- Immune system is completely retrained
Month 3+:
- Stable control of the remaining cancer cells
- Prevents recurrences and metastases
- Regenerate normal cells (especially after chemo)
Dosage recommendation
Dosages of Huaier are NOT generally dependent on body weight and therefore usually do not require adjustment, as the effect is not dependent on blood concentration (like antibiotics, for example), but rather addresses signaling pathways, thereby inducing the intended effects.
The following dosage recommendation results from the above-mentioned studies and applies to all cancers that respond to Huaier (see above).
These dosage recommendations must be adapted to the manufacturer's instructions for Huaier granules! Newer preparations (such as Nutrimentas „MycoPure 58ta-Glucans, Polysaccharides 67.9%“) have an improved formulation and therefore require a 50% reduction to 30 g/d (3 x 10g).
The intake should not be on an empty stomach.
Here is a practical example for metastatic breast cancer after resection and removal of 7 affected lymph nodes, the evidence-based dosage recommendation is based on the Nutrimentas granules with 32% polysaccharides as follows:
Phase 1: Acute phase - after resection Weeks 1-4
Tumor burden: high (7 affected lymph nodes, risk of metastasis)
Recommended total daily amount: 60 g
- Divided: 3 × 20 g daily (morning, noon, evening)
- Time: best on an empty stomach or between meals
Active ingredient content in this phase:
- 60 g × 32% = 19.2 g polysaccharides
- Of which at least: 60 g × 28% = 16.8 g β-glucans
Preparation per dose:
- Pour 20 g of granules into a cup
- Pour approx. 100 ml of hot water (80°C) over it
- Stir well until completely dissolved
- Fill up to approx. 250 ml with lukewarm water
- Drink slowly
Phase 2: Consolidation phase - weeks 5-12
After stabilization and first check-up
Recommended total daily amount: 30 g
- Divided: 3 × 10 g
Active ingredient content in this phase:
- 30 g × 32% = 9.6 g polysaccharides
- Of which at least: 30 g × 28% = 8.4 g β-glucans
This represents the standard dose in oncology and is used in most studies.
Phase 3: Maintenance phase - from the 4th month for a further 6-12 months
Recurrence prevention and metastasis prevention
Recommended total daily amount: 15 g
- 3 × 5 g daily = 15 g
Active ingredient content per day
- 15 g × 32% = 4.8 g polysaccharides
- Of which at least: 15 g × 28% = 4.2 g β-glucans
Important notes:
- Consistency is important: Daily intake without interruption is essential for optimum effect
- Continuous application: In order to ensure the therapeutic effects, the intake should be continued for at least 6-12 months
- Can be combined with conventional medicine: There are no known interactions
- Gentle on the stomach: Better tolerated if the granules are taken on an empty stomach
- Compatibility: Slight detoxification reactions may occur in the first 1-2 weeks (tiredness, headaches). These are normal and subside quickly.
Control examinations
Baseline - before taking Huaier
Blood tests:
- Tumor markers: CEA (carcinoembryonic antigen) - relevant for breast cancer
- Tumor markers: CA 15-3 (particularly important for breast cancer)
- Tumor markers: CA 27.29 (additionally for chest)
- Tumor markers: HER2/new (if not yet known)
- Full blood count: RBC, WBC, Hemoglobin, Hematocrit, Thrombocytes
- Liver function: AST, OLD, GGT, Bilirubin (important, as liver damage is possible with metastases)
- Kidney function: Creatinine, BUN, GFR
- Inflammatory markers: CRP, erythrocyte sedimentation (ISR)
- Immune function: Lymphocyte count (CD4, CD8, NK cells, if possible)
Tumor markers - specific interpretation in breast cancer
CEA (Carcinoembryonic antigen)
- Normal: < 2.5 ng/mL (< 5 ng/mL for smokers)
- What does increase mean? Relapse or metastatic disease
- Sensitivity: 50-70% for metastases
CA 15-3 (Cancer Antigen 15-3)
- Normal: < 25 U/mL (some labs < 35 U/mL)
- What does increase mean? Particularly relevant for metastatic breast cancer
- Sensitivity: 70-80% for metastases, only 25% for early stage
CA 27.29
- Normal: < 38 U/mL
- Which means: Breast cancer-specific marker
- Additional information on CA 15-3
Interpretation under Huaier:
- Good sign: Markers fall continuously or stabilize at a low level
- Warning signal: Continuous increase despite Huaier (= possibly Non-responder*)
- Remark: Individual measured values are not too important, the trends are decisive!
*Recognize non-responders
Warning signals indicate a lack of Huaier efficacy at this dosage:
- Tumor markers rise continuously (despite regular intake of Huaier)
- Lymphocytes remain low (< 20%)
- CT/MRI shows tumor progression
- New metastases on imaging
- Clinical deterioration (weight loss, loss of performance)
In this case, the daily dose of Huaier should be increased to 30-40g/day.
Signs of a positive effect
Blood laboratories:
- ✓ Tumor markers fall continuously
- ✓ Lymphocytes increase
- ✓ Normalization of liver and kidney function
- ✓ CRP (inflammation value) normalizes after initial increase
Imaging:
- ✓ Tumor regression or stabilization
- ✓ Lymph node reduction
- ✓ No new metastases
Clinical condition:
- ✓ Rising energy
- ✓ Improved appetite
- ✓ Better quality of sleep
- ✓ Psychological stabilization
- ✓ Hair growth (signal for stem cell activation)
Blood count parameters
Changes expected while taking Huaier:
Lymphocytes (normal: 20-40% of the WBC)
- Expected change: ↑ increase (= good sign, immune activation)
- Target: > 30%, ideally > 35%
Hemoglobin (normal: 12-16 g/dL in women)
- Expected change: ↑ Stabilization/slight increase
- Huaier supports blood formation (important after chemo)
Platelet count (normal: 150-400 K/μL)
- Expected change: ↑ Stabilization/increase
- Huaier also supports blood formation here
CRP (normal: < 3-5 mg/L)
- Expected change: ↑ Slight increase in week 1-2 (= immune response)
- Then ↓ decline in week 3-4 (= good sign)
- Shows immune activation
Imaging (baseline):
- CT thorax + abdomen (looking for lung metastases and hepatic metastases)
- Skeletal scintigraphy or PET-CT (searches for bone metastases)
- Locoregional assessment (surgical site, axillary lymph nodes)
- Optional: MRI liver (if hepatic involvement is suspected)
Phase 1: Acute phase - weeks 1-4
Dosage: 3 × 20 g daily = 60 g/day
Week 2
- Clinical assessment:
- Tolerance, side effects, energy level
- Appetite, sleep quality
- Gastrointestinal tolerance (nausea, diarrhea)
- Laboratories (optional, only if available):
- Rapid blood count (WBC, Lymphocytes)
- CRP (inflammation)
- Tumor markers (CEA, CA 15-3) - often still too early for significant change
Week 4
- Clinical assessment: General condition, wound healing (if recently operated on)
- Blood tests:
- Tumor markers: CEA, CA 15-3, CA 27.29 (first response check)
- Full blood count (WBC, Lymphocytes, Hemoglobin)
- Liver function (AST, OLD, GGT, Bilirubin)
- Kidney function (Creatinine, GFR)
- CRP (inflammation marker)
- If available: Lymphocyte profile (CD4/CD8 ratio, NK cell count)
- Notes
- ✓ Tumor markers may still rise slightly in this phase (first „detoxification“)
- ✓ Lymphocyte count often increased (immune activation)
- ✓ CRP may be slightly elevated (immune reaction)
Phase 2 - Consolidation phase - weeks 5-12
Reduce dosage to: 3 × 10 g daily = 30 g/day
week 6
- Clinical assessment: Energy level, symptom complaints
Week 8
- Imaging:
- CT thorax + abdomen (first image control)
- Question: Size regression of primary tumor? New metastases? Lymph node regression?
- Comparison with baseline
- Blood tests:
- Tumor markers: CEA, CA 15-3, CA 27.29
- Full blood count
- Liver function
- Kidney function
- Immune markers (if available)
- Notes
- ✓ Tumor markers should now start to fall (or be stable)
- ✓ Imaging should show initial regression or stabilization
- ✓ Lymphocyte count persistently elevated (good sign)
Week 12
- Blood tests:
- Tumor markers (CEA, CA 15-3, CA 27.29)
- Full blood count
- Liver function
- Clinical assessment:
- Decision for phase 3?
- Responder vs. non-responder assessment
Phase 3 - Maintenance phase - from month 4 for 6-12 months
Dosage: 3 × 5 g daily = 15 g/day (or alternatively 2 × 5g = 10g/day)
Month 4 (week 16)
- Blood tests:
- Tumor markers: CEA, CA 15-3, CA 27.29 (response assessment)
- Full blood count
- Liver function, kidney function
- Immune markers
- Clinical assessment:
- Overall evaluation of therapy success to date
- Compatibility, quality of life
- Possible adjustment of dosage based on markers
Month 6 after start
- Imaging (CRITICAL):
- CT thorax + abdomen or PET-CT
- Comparison with week 8 imaging and baseline
- Goal: Confirmation of stable disease or further regression
- Blood tests:
- Tumor markers (CEA, CA 15-3, CA 27.29)
- Full blood count
- Liver function, kidney function
- CRP
- Hormonal markers (if hormone therapy is planned)
Month 9
- Blood tests:
- Tumor markers
- Full blood count
Month 12
- Imaging (FOLLOW-UP):
- CT thorax + abdomen or PET-CT
- Assessment of long-term responses
- Search for delayed metastases
- Blood tests (COMPLETE):
- Tumor markers: CEA, CA 15-3, CA 27.29
- Full blood count
- Liver function, kidney function
- CRP
- Immune markers (if available)
Long-term monitoring from year 2
Dosage: 2 × 3-5 g daily = 6-10 g/day (maintenance)
Every 3 months:
- Blood tests: Tumor markers (CEA, CA 15-3, CA 27.29) + complete blood count
Every 6 months:
- CT or MRI (depending on the oncologist's protocol)
- Complete blood test
Annually:
- Complete baseline examinations (as at the beginning)
- Comprehensive imaging
Function - medically and technically explained
1st Hippo-Pathway
The normal function of the Hippo-Pathway is as follows:
Hippo signaling pathway active
↓
YAP1/TAZ are phosphorylated and inactivated
↓
Transcription of growth genes is stopped
↓
Apoptosis (cellular suicide) or cell cycle arrest
↓
Tumor does not growIn cancer (disturbed Hippo pathway):
Procedure in the event of a disrupted Hippo pathway, e.g. cancer:
Hippo signaling pathway inhibited/mutated
↓
YAP1/TAZ remain active (dephosphorylated)
↓
Uncontrolled transcription of growth genes
↓
Cellular growth is hyperactive
↓
Cancer grows uncontrolledTaking Huaier activates the polysaccharides and metabolites of Huaier:
- LATS1/2 kinases (Upstream regulators of the Hippo pathway)
- This re-phosphorylated YAP1/TAZ
- YAP1/TAZ become again inactivated
- The normal cell cycle control mechanism is restored
2. correction of transcriptional dysregulation
Thousands of genes are switched off in cancer: Genes that should be switched on are switched off and vice versa.
Huaier reactivates the transcription factors:
- NF-κB (Controls immune response and cell survival)
- c-Myc, Oct3/4, Sox2, Klf4 (Pluripotency factors - activate stem cell function)
- p53 (tumor suppressor - induces apoptosis)
- TCF/LEF (Wnt signal pathway effectors)
In addition, mass gene expression is reversed (within 4 weeks according to the Tanaka study)
12,000 to 25,000 new genes (normal cells only have ~20,000 in total) and 8,000 to 15,000 are silenced (switched off)
This leads to a massive „reprogramming“ of the cancer cell:
- Return to stem cell-like properties (non-differentiated)
- Apoptosis pathways are activated
Or: - Differentiation into the normal cell type (cell specialization) takes place
Through the reactivation of stem cell genes (c-myc, Oct3/4), the cancer cell becomes sensitive to normal control mechanisms again.
3. PI3K/AKT/mTOR signal path modulation
Normal (inhibited):
PI3K active → AKT active → mTOR active → Cell growth inhibited ✓
(This is oversimplified, but the concept)For cancer (hyperactive):
PI3K overactive → AKT overactive → mTOR hyperactive → Uncontrolled growth ✗
(This is one of the most common defects in cancer cells)Huaier has the effect that
- PTEN activated (negative regulator of PI3K)
- TSC1/TSC2 complexes become restored (inhibit mTOR)
- PI3K/AKT/mTOR is converted into normal Balance returned
- Cell growth becomes controllable again
Note: This route is particularly suitable for HER2-negative and Triple negatives Breast cancer excessively active.
4. miRNA- and piRNA-mediated transcriptional control
MicroRNAs (miRNA, small pieces of RNA (molecules) with a length of 20-22 nucleotides) are normally the „brakes“ for faulty genes. In cancer, these brakes are disrupted:
- Oncogenes are no longer inhibited
- Tumor suppressor genes are slowed down too much
Huaier provides the Restoration of miRNA function:
- miR-122 (inhibits HCC growth)
- miR-145 (inhibits stem cell genes in normal cells)
- miR-17/92 cluster (is activated by c-myc, can then induce apoptosis)
New miRNAs are activated, which:
- Oncogenes (e.g. KRAS, PIK3CA) shut down
- Tumor suppressor genes (TP53, RB) reinforce
- Angiogenesis (blood vessel formation) inhibit
- Epithelial-Mesenchymal Transition (EMT) block → Blocks metastasis
Tanaka study: Hundreds of new miRNA-variants that specifically „mute“ cancer cells.
5. immune activation (innate immune system)
β-Glucans as pattern recognition ligands:
β-Glucans (from Huaier)
↓
Binding to Dectin-1 and TLR receptors on immune cells
↓
Activation of macrophages and NK cells
↓
Secretion of pro-inflammatory cytokines:
- TNF-α (tumor necrosis factor)
- IL-12 (interleukin-12)
- IFN-γ (interferon-gamma)
↓
Activation of cytotoxic T cells (CD8+)
↓
Recognition and lysis of tumor cellsThe immune system is virtually „shaken awake“ and recognizes cancer cells as enemies again.
6. blockade of epithelial-mesenchymal transition (EMT)
The EMT process causes cancer cells to lose adhesion to their base, allowing them to migrate around the organism and lead to metastasis:
- Cells lose E-Cadherin (cellular adhesive)
- Cells express vimentin (movement protein)
Huaier provides the
- Stabilization from E-Cadherin (cells „stick“ together again)
- curtailment from Vimentin (cells can „migrate“ less)
- inhibition from Snail-, Slug- and Twist factors (EMT inducers)
- Stabilization from β-Catenin (maintains normal epithelial function)
This mechanically blocks the formation of metastases, even in the case of existing lymph node metastases.
7. blockage of angiogenesis (blood vessel formation)
Tumors can only grow if they form new blood vessels (angiogenesis). This is driven by VEGF (vascular endothelial growth factor).
Huaier counteracts this by
- VEGF expression inhibited becomes
- VEGFR signaling pathways blocked become
- HIF-1α (hypoxia inducible factor) downregulated becomes
- alternative pro-angiogenic pathways (FGF, Notch) inhibited become
Result: The tumor loses its blood supply - tumor growth is inhibited.
8. ribosomal RNA structure repair
The problem after chemotherapy:
- Chemotherapeutic agents, especially platinum complexes such as cisplatin, destroy ribosomal RNA structures
- Ribosomes are the protein factories of the cell
- Without functional ribosomes, the cell cannot produce proteins
Even if the tumor dies, healthy cells cannot regenerate
Huaier intervenes here and
- repairs ribosomal RNA structures
- provides the Restore protein synthesis capacity
This allows healthy cells to regenerate, while cancer cells die again. This explains why Huaier patients undergoing chemotherapy have fewer side effects and recover more quickly.
Immunologically relevant genes
Regulatory behavior of genes
Genes can be linked to 0% Expression (corresponds practically to OFF) or with any percentage of their maximum capacity, or with 100% Expression (for complete AN).
Tumor necrosis factor α
The regulatory behavior and its consequences are explained using the example of the tumor necrosis factor α (TNFα):
The normal “value“ is 40% expression, enough to protect against infections, too little to attack tissue.
If the value rises to 100% (or even higher), e.g. in rheumatoid arthritis, this results in
- a massive excess of TNF-α
- Permanent joint inflammation
- Destruction of cartilage and bone
- systemic inflammation
and the symptoms of permanent joint pain and swelling.
If, however, the value is reduced to only 5%, for example, then this results in
- Too little TNF-α to kill pathogens
- Unlimited bacterial growth
- Systemic organ failure
- Fatalities possible
Conclusion: TNF is vital!
Spectrum of the cytokine IL-6 (interleukin 6)
- Silent - 0-5% of the control value
No acute phase reaction, no fever
Infection blindness - Very quiet - 5-15% of the control value
Minimal inflammatory response
Weak immunity - Quiet - 15-30% of the control value
Mild local inflammation
NORMAL after a minor infection - Moderate - 30-50% of the control value
Clear but limited inflammation
NORMAL in case of infection - Loud - 50-80% of the control value
Severe systemic inflammation
Too much? For RA, IBD - Very loud - 80-95% of the control value
Massive systemic inflammation
Sepsis, shock - Maximum - 95-100%+ of the control value
Cytokine storm, organ failure
Fatal (COVID-19, sepsis)
Example of low expression
- TNF-α at 90% instead of 40% of the control value
Autoimmune inflammation - IL-17 at 85% instead of 30% of the control value
Overproduction of Th17 leads to excessive inflammatory reactions - IL-6 at 95% instead of 45% of the control value
Chronic arthritis
Example of excessive expression
- TNF-α at 10% instead of 40% of the control value
Risk of tuberculosis - IL-10 at 8% instead of 35% of the control value
Inflammation uncontrolled - IFN-γ at 12% instead of 50% of the control value
Viral susceptibility
Measurement methods
The regulation of genes takes place on several biological levels. To understand these levels, there are four main measurement methods that quantify different aspects of gene expression:
- Level 1: Transcription (DNA → mRNA)
Measurement method: qRT-PCR - Protein production level (mRNA → protein in the cell)
Measurement method: Western Blotting - Level 3: Secretion/circulation (protein in serum/plasma)
Measurement method: ELISA - Level 4: Cellular expression at single cell level
Measurement method: Flow Cytometry
1. qRT-PCR (Quantitative Reverse Transcription PCR)
qRT-PCR measures the amount of mRNA in cells or tissues in the measurand „multiple“
- Fold-Change (multiple): Example: TNF-α mRNA is increased 2.5-fold
- Significance: 2.5× higher than the control group
- A value of 0.45 means: 45% of the control (i.e. down-regulated)
- Cycle Threshold (Ct): Raw value, how many PCR cycles are required until detection
- Lower Ct = more mRNA present
- Higher Ct = less mRNA present
What qRT-PCR does NOT measure:
- the absolute amount of protein
- the activity of the protein
- whether the protein has been secreted
- the concentration in the serum
Clinical interpretation
qRT-PCR: TNF-α = 2.5-fold
Means: "TNF-α mRNA is 2.5× higher than normal"
"The gene 'volume control' is turned up"
BUT: This says NOTHING about the actual amount of TNF-α protein in the serum!
Flow cytometry shows that even high mRNA does not automatically produce a lot of protein per cell, and even if it does, it still has to be secreted. The 8.3-fold mRNA increase in the following example can therefore lead to more or less protein in the cells.Practical example
Patient with bacterial infection:
qRT-PCR (blood leukocytes): TNF-α = 8.3-fold increased
→ The cells produce a lot of mRNA
→ but this is not immediately measurable in the serum
→ because the protein only arrives in the serum after about 30 minutes to hours.2. western blot
The Western Blot measures the amount of protein within cells or tissues in the „band intensity“ measure, the phosphorylation status (activated vs. inactive protein) and various protein isoforms.
- Relative band intensity: 0-100% or as a multiple for control purposes
- Example: IL-6 protein = 65% of the control intensity
- Significance: The protein is as strongly expressed at 65% as in the control
What Western Blotting does NOT measure:
- whether the protein is active (presence only)
- whether the protein has been secreted
- the concentration in the serum/blood
- at single cell level
Clinical interpretation:
Western blot: TNF-α protein = 72% of the control intensity
Means: "TNF-α protein is present at 72% in the cell lysate"
"72% as much protein as in the control cell culture"
BUT: This says NOTHING about:
- How much TNF-α was actually secreted
- How much TNF-α is in the serum
- Whether the protein is active or notPractical example:
Macrophage culture with LPS stimulation:
Western blotting (cell lysate): TNF-α = 85% of the control
ELISA (culture supernatant): TNF-α = 2,800 pg/mL
Conclusion: A lot of TNF-α protein was produced AND secreted3. enzyme-linked immunosorbent assay (ELISA)
ELISA measures the absolute concentration of protein in serum, plasma, cell culture supernatant or other body fluids in absolute concentration.
- pg/mL (picogram per milliliter)
for cytokines such as TNF-α, IL-6 - ng/mL (nanogram per milliliter)
for more concentrated proteins - µg/mL (micrograms per milliliter)
for very high concentrations
Normal values for TNF-α (example):
Healthy: 5,000 pg/mL (can be fatal)What ELISA measures:
- Absolute amount of secreted/circulating protein
- whether the protein has actually arrived in the blood/serum
- the systemic effect (not just locally in the cell)
What ELISA does NOT measure:
- how much mRNA is present
- how much protein is in the cells
- whether the protein is active
- at single cell level
Clinical interpretation:
ELISA: TNF-α in serum = 65 pg/mL
Means: "There are 65 picograms of TNF-α per milliliter of serum"
"This is 3-13× above the normal value"
"There is moderate inflammation"
This is an ABSOLUTE concentration, not relative!Practical example:
Patient with rheumatoid arthritis:
ELISA: TNF-α = 85 pg/mL (normal: < 20 pg/mL)
qRT-PCR (blood): TNF-α mRNA = 3.2-fold increased
Western blotting (joint synovia): TNF-α = 95% (very high locally)
Conclusion: Too much TNF-α everywhere, from mRNA to cellular protein to serum4. flow cytometry
Flow cytometry measures the expression of proteins or markers on the surface or inside individual cells as a percentage and fluorescence intensity.
- % positive cells: Example: 78% of CD4+ T cells express IL-2
- Significance: 78% of this cell population has the characteristic
- Mean Fluorescence Intensity (MFI): 0-10,000+ (depending on instrument)
- Example: IL-2 expression MFI = 450 in CD4+ T cells
- Higher MFI = more protein per cell
What Flow Cytometry measures:
- How many cells of a certain population express an antigen (%)
- How much antigen is present per cell (MFI)
- Cellular heterogeneity (not all cells are the same!)
- Cell surface markers and intracellular proteins
What Flow Cytometry does NOT measure:
- The serum concentration (measures cells, not serum)
- The amount of mRNA - How much is in the body in total -
Additional data (cell count, weight, etc.) can be used to make indirect extrapolations:
However, this extrapolation is only an estimate, not as accurate as ELISA
and it only records the measured cells (e.g. blood macrophages), not tissue macrophages!
Clinical interpretation:
Flow cytometry: 73% of CD8+ T cells express IFN-γ
MFI = 520
Means: "73% of cytotoxic T cells have IFN-γ protein"
"The average IFN-γ content per cell is 520 (MFI)"
"The T cell response is active"
BUT: This says NOTHING about:
- How much total IFN-γ is in the serum
- How much IFN-γ mRNA is presentPractical example:
COVID-19 patient (day 3 after infection):
Flow cytometry:
- 91% CD8+ T cells express IFN-γ (high!)
- MFI = 1.250 (very high)
ELISA: IFN-γ in serum = 180 pg/mL (normal: < 50)
Conclusion: Strong T-cell-activated IFN-γ production, systemically measurable