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Create physiologically relevant cell culture models

Gibco Human Plasma-like Medium (HPLM) is a new formulation designed to resemble the natural cellular environment found in the body, mimicking the metabolic profile of human plasma. 

The widely used classic synthetic cell culture media, including MEM, DMEM, RPMI 1640, and DMEM/F-12 contain glucose, amino acids, vitamins, and salts at concentrations that in large part do not reflect those found in human plasma. These media also lack additional plasma components needed to mimic the metabolic profile of human plasma. When studying cancer and other diseases, results with more physiological relevance will enable researchers to improve their understanding of human function and illness.

The HPLM solution

Gibco HPLM contains the same salt concentrations found in human plasma, as well as the same concentrations of over 60 polar metabolites, such as amino acids, nucleic acids, sugars, and small organic acids. In resembling the natural cellular environment found in the body, HPLM provides researchers the ability to study the impact of physiologically relevant cell media on their specific applications.

When supplemented with fetal bovine serum (FBS), HPLM is capable of supporting cell growth and viability that are comparable to that of conventional basal media formulations supplemented with FBS. For most cell lines, adaptation is not required to transition from conventional medium to HPLM.

  • Physiologically relevant—designed to resemble the natural cellular environment found in the body formulated with more than 60 polar metabolites and salt concentrations
  • Peer reviewed—extensive research publications completed using HPLM formulation
  • Easy to use—direct replacement for your current media when supplemented with FBS

Order now   View data   Explore publications


Meet the inventors

Headshot of Jason R. Cantor

 

Headshot of David M. Sabatini

 


The inventors of human plasma-like medium are Jason R. Cantor and David M. Sabatini.

As a postdoc in the laboratory of David M. Sabatini at the Whitehead Institute/MIT in Cambridge, Jason set out to create what would become human plasma-like medium (HPLM), a physiologic medium designed to more closely reflect the metabolic composition of human blood, thus permitting the study of cultured cells in biochemical conditions with greater relevance to human physiology.

Cantor and Sabatini ultimately reported their development and initial studies using HPLM in early 2017 (Cell). Read their publication here, Physiologic Medium Rewires Cellular Metabolism and Reveals Uric Acid as an Endogenous Inhibitor of UMP Synthase.

We are proud to work with Jason and David to bring this innovation to market, and excited by the immense possibilities that HPLM could bring across diverse areas of the scientific community. As Jason notes, "The recent development of physiologic media, like other efforts designed to address the modeling capacity of cell culture, holds immense potential to improve understanding and interpretation of diverse biological and pharmacological studies." Read more from his 2019 commentary here: The Rise of Physiologic Media.

Jason and David are inventors on a patent application for HPLM assigned to Whitehead Institute.


Research results using human plasma-like medium

HPLM rewires cellular metabolism

Graphical abstract summary: “Among the most prominent was an inhibition of de novo pyrimidine synthesis—an effect traced to uric acid, which is 10-fold higher in the blood of humans than of mice and other non-primates. We find that uric acid directly inhibits uridine monophosphate synthase (UMPS) and consequently reduces the sensitivity of cancer cells to the chemotherapeutic agent 5-fluorouracil. Thus, media that better recapitulates the composition of human plasma reveals unforeseen metabolic wiring and regulation, suggesting that HPLM should be of broad utility.”

Reproduced with permission from: Cantor JR, Abu-Remaileh M, Kanarkek N et al. (2017) Physiologic medium rewires cellular metabolism and reveals uric acid as an endogenous inhibitor of UMP synthase. Cell 169: 258–272.E17. doi: 10.1016/j.cell.2017.03.023.

HPLM improves T lymphocyte activation

Graphical abstract summary: “The composition of human plasma differs from conventional media, and we hypothesized that such differences could impact immune cell physiology. Here, we showed that relative to the medium typically used to culture lymphocytes (RPMI), a physiologic medium (human plasma-like medium; HPLM) induced markedly different transcriptional responses in human primary T cells and in addition, improved their activation upon antigen stimulation. We found that this medium-dependent effect on T cell activation is linked to Ca2+, which is six-fold higher in HPLM than in RPMI.  Thus, a medium that more closely resembles human plasma has striking effects on T cell biology, further demonstrates that medium composition can profoundly affect experimental results, and broadly suggests that physiologic media may offer a valuable way to study cultured immune cells."

Reproduced with permission from: Leney-Greene MA, Boddapati AK, Su HC et al. (2020) Human plasma-like medium improves T lymphocyte activation. iScience 23:100759. doi: 10.1016/j.isci.2019.100759.

Human Plasma-Like Medium maintains cell morphology

2-panel brightfield microscopic views of MCF7 cells growing in culture in DMEM and HPLM.

Figure 1. Gibco HPLM supports MCF7 cell culture. MCF7 breast adenocarcinoma cells were cultured in DMEM (Cat. No. 10566016) or Human Plasma-Like Medium (HPLM, Cat. No. A4899101) supplemented with 10% FBS (Cat. No. A3840101).

2-panel brightfield microscopic views of HeLa cells growing in culture in DMEM and HPLM.

Figure 2. Gibco HPLM supports HeLa cell culture. HeLa human cervical adenocarcinoma cells were cultured in DMEM (Cat. No. 11965092) or Human Plasma-Like Medium (HPLM, Cat. No. A4899101) supplemented with 10% FBS (Cat. No. A3840101).

2-panel brightfield microscopic views of LNCaP cells growing in culture in DMEM and HPLM.

Figure 3. Gibco HPLM supports LNCaP cell culture. LNCaP human metastatic prostate carcinoma cells were cultured in RPMI 1640 (left; Cat. No. 61870036) or HPLM (right; Cat. No. A4899101) supplemented with 10% FBS (Cat. No. A3840101).

Human Plasma-Like Medium maintains cell growth

4-panel bar charts showing doubling times for 4 cell lines in DMEM and in HPLM.
Figure 4. Gibco HPLM supports comparable growth rates in continuous culture. Tumor cell lines A549, HeLa, MCF7, and THP-1 were grown in DMEM (blue) or HPLM (red) supplemented with 10% FBS (Cat. No. A3840101) for 5 passages. Cell number was assessed at the end of each passage and used to calculate the average population doubling time for each culture.


Human Plasma-like Medium supports cell growth in multiple cell types

Table 1. Cell types successfully grown in Human Plasma-like Medium.

Cell type tested Cell origin
HeLa, adherent Cervical adenocarcinoma
A549, adherent Lung carcinoma
MCF-7, adherent Metastatic breast cancer
MDA-MB-231, adherent Metastatic breast cancer
LNCaP, adherent Metastatic prostate carcinoma
HCT116, adherent Colorectal carcinoma
A375, adherent Malignant melanoma
WM115, adherent Malignant melanoma
U-2 OS, adherent Osteosarcoma
NK, primary, suspension Lymphocyte, blood
THP-1, suspension Monocytic leukemia
Jurkat, suspension T cell leukemia
PLB-985, suspension Myeloid leukemia
NOMO-1, suspension Monoblastic/monocytic leukemia
Sp2, suspension Mouse B lymphocyte

The following customer stories feature the work of our Gibco HPLM product testers.

Headshot of Omar Torres-Quesada
“We used the HPLM media with cancer cell lines from different tissues and different mutation background. Our overall impression of HPLM is quite positive. HPLM media was suitable for all the tested cancer cell lines. Cells were easily and quickly adapted to the HPLM without affecting viability and cell culture performance. However, we detected relevant impact of cell proliferation, metabolism and mitochondrial function in cells grown in HPLM as compared with classical media. After our experience using HPLM in cancer cell models we consider it as the best choice to get more physiological data.”

— Omar Torres-Quesada, PhD, Postdoctoral Researcher at University of Innsbruck, Austria

Headshot of Justin Rettenmaier
“Immunometabolism is an exciting area of scientific investigation that has immense potential for the development of new therapeutics. It is becoming clear that how immune cells rewire their metabolism after activation depends on what nutrients are available and in what quantities. Since we study metabolite transport in primary human immune cells, we find HPLM media to be essential for modeling metabolic flux in the most physiologically relevant way possible. We have extensively vetted the use of HPLM across CD4 T helper cell lineages and find that it supports both proliferation and effector function."

— Justin Rettenmaier, Associate Director at Jnana Therapeutics, Boston, MA

Headshot of Martin Fischer
"Standard cell culture media, including the Dulbecco’s Modified Eagle Medium (DMEM), contain a non-physiological excess of nutrients, including glucose and glutamine. At the same time, they are low on uric acid. Nutrient availability, however, is a most critical factor for the regulation of mTOR calling for a careful consideration of culture conditions. Therefore, we employed Human Plasma Like Medium (HPLM) to gain a better understanding of mTORC1 regulation under physiological nutrient abundance. In line with mTOR’s intricate connection to nutrient sensing pathways, mTORC1 activity, as measured by Thr389 phosphorylation of S6K, was lower when U2OS cells were cultured in HPLM instead of DMEM. In HPLM media and irrespective of any treatment, AKT and the glucose-sensing AMPK were activated. Most importantly, however, we found that p53 was required to sustain low mTORC1 activity under HPLM culture conditions irrespective of Nutlin-3a treatment. Together, our investigation using HPLM revealed a nutrient-dependent role of p53 in mTORC1 inhibition."

— Martin Fischer, Principal Investigator at Leibniz Institute on Aging, Germany

HPLM rewires cellular metabolism

Graphical abstract summary: “Among the most prominent was an inhibition of de novo pyrimidine synthesis—an effect traced to uric acid, which is 10-fold higher in the blood of humans than of mice and other non-primates. We find that uric acid directly inhibits uridine monophosphate synthase (UMPS) and consequently reduces the sensitivity of cancer cells to the chemotherapeutic agent 5-fluorouracil. Thus, media that better recapitulates the composition of human plasma reveals unforeseen metabolic wiring and regulation, suggesting that HPLM should be of broad utility.”

Reproduced with permission from: Cantor JR, Abu-Remaileh M, Kanarkek N et al. (2017) Physiologic medium rewires cellular metabolism and reveals uric acid as an endogenous inhibitor of UMP synthase. Cell 169: 258–272.E17. doi: 10.1016/j.cell.2017.03.023.

HPLM improves T lymphocyte activation

Graphical abstract summary: “The composition of human plasma differs from conventional media, and we hypothesized that such differences could impact immune cell physiology. Here, we showed that relative to the medium typically used to culture lymphocytes (RPMI), a physiologic medium (human plasma-like medium; HPLM) induced markedly different transcriptional responses in human primary T cells and in addition, improved their activation upon antigen stimulation. We found that this medium-dependent effect on T cell activation is linked to Ca2+, which is six-fold higher in HPLM than in RPMI.  Thus, a medium that more closely resembles human plasma has striking effects on T cell biology, further demonstrates that medium composition can profoundly affect experimental results, and broadly suggests that physiologic media may offer a valuable way to study cultured immune cells."

Reproduced with permission from: Leney-Greene MA, Boddapati AK, Su HC et al. (2020) Human plasma-like medium improves T lymphocyte activation. iScience 23:100759. doi: 10.1016/j.isci.2019.100759.

Human Plasma-Like Medium maintains cell morphology

2-panel brightfield microscopic views of MCF7 cells growing in culture in DMEM and HPLM.

Figure 1. Gibco HPLM supports MCF7 cell culture. MCF7 breast adenocarcinoma cells were cultured in DMEM (Cat. No. 10566016) or Human Plasma-Like Medium (HPLM, Cat. No. A4899101) supplemented with 10% FBS (Cat. No. A3840101).

2-panel brightfield microscopic views of HeLa cells growing in culture in DMEM and HPLM.

Figure 2. Gibco HPLM supports HeLa cell culture. HeLa human cervical adenocarcinoma cells were cultured in DMEM (Cat. No. 11965092) or Human Plasma-Like Medium (HPLM, Cat. No. A4899101) supplemented with 10% FBS (Cat. No. A3840101).

2-panel brightfield microscopic views of LNCaP cells growing in culture in DMEM and HPLM.

Figure 3. Gibco HPLM supports LNCaP cell culture. LNCaP human metastatic prostate carcinoma cells were cultured in RPMI 1640 (left; Cat. No. 61870036) or HPLM (right; Cat. No. A4899101) supplemented with 10% FBS (Cat. No. A3840101).

Human Plasma-Like Medium maintains cell growth

4-panel bar charts showing doubling times for 4 cell lines in DMEM and in HPLM.
Figure 4. Gibco HPLM supports comparable growth rates in continuous culture. Tumor cell lines A549, HeLa, MCF7, and THP-1 were grown in DMEM (blue) or HPLM (red) supplemented with 10% FBS (Cat. No. A3840101) for 5 passages. Cell number was assessed at the end of each passage and used to calculate the average population doubling time for each culture.


Human Plasma-like Medium supports cell growth in multiple cell types

Table 1. Cell types successfully grown in Human Plasma-like Medium.

Cell type tested Cell origin
HeLa, adherent Cervical adenocarcinoma
A549, adherent Lung carcinoma
MCF-7, adherent Metastatic breast cancer
MDA-MB-231, adherent Metastatic breast cancer
LNCaP, adherent Metastatic prostate carcinoma
HCT116, adherent Colorectal carcinoma
A375, adherent Malignant melanoma
WM115, adherent Malignant melanoma
U-2 OS, adherent Osteosarcoma
NK, primary, suspension Lymphocyte, blood
THP-1, suspension Monocytic leukemia
Jurkat, suspension T cell leukemia
PLB-985, suspension Myeloid leukemia
NOMO-1, suspension Monoblastic/monocytic leukemia
Sp2, suspension Mouse B lymphocyte

The following customer stories feature the work of our Gibco HPLM product testers.

Headshot of Omar Torres-Quesada
“We used the HPLM media with cancer cell lines from different tissues and different mutation background. Our overall impression of HPLM is quite positive. HPLM media was suitable for all the tested cancer cell lines. Cells were easily and quickly adapted to the HPLM without affecting viability and cell culture performance. However, we detected relevant impact of cell proliferation, metabolism and mitochondrial function in cells grown in HPLM as compared with classical media. After our experience using HPLM in cancer cell models we consider it as the best choice to get more physiological data.”

— Omar Torres-Quesada, PhD, Postdoctoral Researcher at University of Innsbruck, Austria

Headshot of Justin Rettenmaier
“Immunometabolism is an exciting area of scientific investigation that has immense potential for the development of new therapeutics. It is becoming clear that how immune cells rewire their metabolism after activation depends on what nutrients are available and in what quantities. Since we study metabolite transport in primary human immune cells, we find HPLM media to be essential for modeling metabolic flux in the most physiologically relevant way possible. We have extensively vetted the use of HPLM across CD4 T helper cell lineages and find that it supports both proliferation and effector function."

— Justin Rettenmaier, Associate Director at Jnana Therapeutics, Boston, MA

Headshot of Martin Fischer
"Standard cell culture media, including the Dulbecco’s Modified Eagle Medium (DMEM), contain a non-physiological excess of nutrients, including glucose and glutamine. At the same time, they are low on uric acid. Nutrient availability, however, is a most critical factor for the regulation of mTOR calling for a careful consideration of culture conditions. Therefore, we employed Human Plasma Like Medium (HPLM) to gain a better understanding of mTORC1 regulation under physiological nutrient abundance. In line with mTOR’s intricate connection to nutrient sensing pathways, mTORC1 activity, as measured by Thr389 phosphorylation of S6K, was lower when U2OS cells were cultured in HPLM instead of DMEM. In HPLM media and irrespective of any treatment, AKT and the glucose-sensing AMPK were activated. Most importantly, however, we found that p53 was required to sustain low mTORC1 activity under HPLM culture conditions irrespective of Nutlin-3a treatment. Together, our investigation using HPLM revealed a nutrient-dependent role of p53 in mTORC1 inhibition."

— Martin Fischer, Principal Investigator at Leibniz Institute on Aging, Germany

Recent publications using human plasma-like medium

Human Plasma-like Medium Improves T Lymphocyte Activation
Leney-Greene MA, Boddapati AK, Su HC, Cantor JR, Lenardo MJ. iScience. 2020;23(1):100759.

  • Research area: Immunology 
  • Cell lines: Peripheral blood mononuclear cells (PBMCs), T lymphocytes
  • Objective: Test the impact of HPLM versus RPMI on T-lymphocyte activation
  • Findings: “Our data broadly highlight the fact that commonly used conditions used to culture and examine T lymphocytes in vitro may not be ideal for metabolic studies, and we also specifically identify one extracellular component (calcium) that can be easily considered by others in the field. Our approach also further demonstrates the value of using HPLM to improve the modeling capacity of in vitro cell culture systems.”

Lineage-Specific Silencing of PSAT1 Induces Serine Auxotrophy and Sensitivity to Dietary Serine Starvation in Luminal Breast Tumors
Choi BH, Conger KO, Selfors LM, Coloff JL. bioRxiv 2020.06.19.161844.

  • Research area: Cancer therapy, gene expression metabolism
  • Cell lines and culture conditions: Luminal and basal human breast tumors:
    • HCC1 806, MCF7, MCF7-EMPTY, MCF7-PSAT1
    • Cells were grown in human plasma-like medium according to the published formulation (Cantor et al., 2017) with 5% dialyzed FBS. Media was changed at least every two days.
  • Objective: Identify differences in metabolic gene expression that may limit pathway redundancy and create therapeutic vulnerabilities
  • Findings: “In summary, our studies of lineage-dependent gene expression in breast cancer have revealed a novel vulnerability in serine metabolism specifically in luminal breast tumors. Lineage-specific suppression of PSAT1 induces serine auxotrophy in luminal breast cancer cells and sensitizes them serine starvation. These findings demonstrate that lineage-dependent gene 10 expression is sufficient to limit pathway redundancy and create therapeutic vulnerabilities that could be taken advantage of to target specific subtypes of tumors.”
     

Inhibiting both proline biosynthesis and lipogenesis synergistically suppresses tumor growth
Liu M, Wang Y, Yang C, Ruan Y, Bai C, Chu Q, Cui Y, Chen C, Ying G, Li B. J Exp Med. 2020 Mar 2;217(3):e20191226.

  • Research area: Cancer metabolism
  • Cell lines: HeLa, MDA-MB-231, MCF-7, A549, HepG2, 8133, SKOV3, and mouse 4T1 cells
  • Objectives:
    • To build up an electron balance model to reveal the chemical mechanism of metabolic reprogramming under hypoxia
    • HPLM was used to test cellular metabolism at physiological (5%), hypoxic (0.5%), and normal oxygen (20%) concentrations
  • Findings: “Overall, our model suggests a promising combination of targets to control tumor growth under hypoxia. Moreover, this treatment can be further intensified by blocking electron transfer established on our concept. Therefore, it is anticipated that the model of electron balance developed in the current study could help us to better understand cancer metabolism and its therapeutic application in the future.”
     

CRISPR screens in physiologic medium reveal conditionally essential genes in human cells
Rossiter NJ, Huggler KS, Adelmann CH, Keys HR, Soens RW, Sabatini DM, Cantor JR.Cell Metab.2021;S1550-4131(21)00061-9.

  • Research type: Cancer biology
  • Cell lines: K562, MOLM-13, SUDHL4, and NOMO1
  • Objective: To explore how medium composition influences gene essentiality of human cancer cell lines cultured in traditional medium versus HPLM
  • Findings: “To explore how medium composition influences gene essentiality, we performed CRISPR-based screens of human cancer cell lines cultured in traditional versus human plasma-like medium (HPLM). Sets of medium-dependent fitness genes span several cellular processes and can vary with both natural cell-intrinsic diversity and the specific combination of basal and serum components that comprise typical culture media. Our findings reveal the profound impact of medium composition on gene essentiality in human cells, and also suggest general strategies for using genetic screens in HPLM to uncover new cancer vulnerabilities and gene-nutrient interactions.”

Human Plasma-like Medium Improves T Lymphocyte Activation
Leney-Greene MA, Boddapati AK, Su HC, Cantor JR, Lenardo MJ. iScience. 2020;23(1):100759.

  • Research area: Immunology 
  • Cell lines: Peripheral blood mononuclear cells (PBMCs), T lymphocytes
  • Objective: Test the impact of HPLM versus RPMI on T-lymphocyte activation
  • Findings: “Our data broadly highlight the fact that commonly used conditions used to culture and examine T lymphocytes in vitro may not be ideal for metabolic studies, and we also specifically identify one extracellular component (calcium) that can be easily considered by others in the field. Our approach also further demonstrates the value of using HPLM to improve the modeling capacity of in vitro cell culture systems.”

Lineage-Specific Silencing of PSAT1 Induces Serine Auxotrophy and Sensitivity to Dietary Serine Starvation in Luminal Breast Tumors
Choi BH, Conger KO, Selfors LM, Coloff JL. bioRxiv 2020.06.19.161844.

  • Research area: Cancer therapy, gene expression metabolism
  • Cell lines and culture conditions: Luminal and basal human breast tumors:
    • HCC1 806, MCF7, MCF7-EMPTY, MCF7-PSAT1
    • Cells were grown in human plasma-like medium according to the published formulation (Cantor et al., 2017) with 5% dialyzed FBS. Media was changed at least every two days.
  • Objective: Identify differences in metabolic gene expression that may limit pathway redundancy and create therapeutic vulnerabilities
  • Findings: “In summary, our studies of lineage-dependent gene expression in breast cancer have revealed a novel vulnerability in serine metabolism specifically in luminal breast tumors. Lineage-specific suppression of PSAT1 induces serine auxotrophy in luminal breast cancer cells and sensitizes them serine starvation. These findings demonstrate that lineage-dependent gene 10 expression is sufficient to limit pathway redundancy and create therapeutic vulnerabilities that could be taken advantage of to target specific subtypes of tumors.”
     

Inhibiting both proline biosynthesis and lipogenesis synergistically suppresses tumor growth
Liu M, Wang Y, Yang C, Ruan Y, Bai C, Chu Q, Cui Y, Chen C, Ying G, Li B. J Exp Med. 2020 Mar 2;217(3):e20191226.

  • Research area: Cancer metabolism
  • Cell lines: HeLa, MDA-MB-231, MCF-7, A549, HepG2, 8133, SKOV3, and mouse 4T1 cells
  • Objectives:
    • To build up an electron balance model to reveal the chemical mechanism of metabolic reprogramming under hypoxia
    • HPLM was used to test cellular metabolism at physiological (5%), hypoxic (0.5%), and normal oxygen (20%) concentrations
  • Findings: “Overall, our model suggests a promising combination of targets to control tumor growth under hypoxia. Moreover, this treatment can be further intensified by blocking electron transfer established on our concept. Therefore, it is anticipated that the model of electron balance developed in the current study could help us to better understand cancer metabolism and its therapeutic application in the future.”
     

CRISPR screens in physiologic medium reveal conditionally essential genes in human cells
Rossiter NJ, Huggler KS, Adelmann CH, Keys HR, Soens RW, Sabatini DM, Cantor JR.Cell Metab.2021;S1550-4131(21)00061-9.

  • Research type: Cancer biology
  • Cell lines: K562, MOLM-13, SUDHL4, and NOMO1
  • Objective: To explore how medium composition influences gene essentiality of human cancer cell lines cultured in traditional medium versus HPLM
  • Findings: “To explore how medium composition influences gene essentiality, we performed CRISPR-based screens of human cancer cell lines cultured in traditional versus human plasma-like medium (HPLM). Sets of medium-dependent fitness genes span several cellular processes and can vary with both natural cell-intrinsic diversity and the specific combination of basal and serum components that comprise typical culture media. Our findings reveal the profound impact of medium composition on gene essentiality in human cells, and also suggest general strategies for using genetic screens in HPLM to uncover new cancer vulnerabilities and gene-nutrient interactions.”

Ordering information

Item Cat No.
2-hydroxybutyric acid (available through Alfa Aesar) Alfa Aesar A18636-03
Fetal Bovine Serum, dialyzed, US origin 26400044
6 Well Plate, TC Surface, Pack of 1 140675
96 Well Plate, TC Surface, Pack of 1 167008
150mm EasYDish, TC Surface, Pack of 10 150468
DPBS, no calcium, no magnesium 14190144
TrypLE Express Enzyme (1X), no phenol red 12604013
Trypsin-EDTA, 0.05%, phenol red 25300054
Trypan Blue Solution, 0.4% 15250061
Countess 3 Automated Cell Counter AMQAX2000

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