Pick Preferred Biospecimen to Perfect Immune Profiling
Simple, underappreciated solutions for reliable flow cytometry data and longitudinal study design
Complex problems call for custom, creative solutions. Immune profiling is essential for developing many therapeutics, including cell and gene therapies. Think about how fast mRNA therapeutics and vaccines were developed to treat and slow COVID-19. Clinical and translational researchers ran countless immune profiling experiments on biospecimens. Those experiments helped build a detailed picture of how the SARS-CoV-2 virus, the immune system, and various treatments interact.
Immune profiling has grown fast over the last decade. You can see it in the rising number of publications. You can also see it in the $180 million the National Institutes of Health (NIH) set aside for immune profiling studies in 2020 [Pushparaj 2020].
Careful selection and handling of biospecimens is critical to the success of these studies.
The Gold Standard of Immune Profiling
Researchers have several techniques for profiling, characterizing, and studying different parts of the immune system and how they relate to each other. Common techniques include:
- Next-generation sequencing (bulk and single-cell approaches)
- Multiplex assays
- Flow cytometry
Flow cytometry may be the gold standard of immune profiling. It can identify, sort, and quantitatively characterize subsets of fluorescently labeled immune cells within a mixed cell population — a capability few other techniques match.
These strengths have made flow cytometry a critical tool in discovery, translational, and clinical biomarker research. It’s also essential for checking the quality of cell & gene therapy products. As flow cytometry keeps spreading across the drug development pipeline, its market is projected to grow from $6.2 billion in 2022 to $16.6 billion by 2032 [Shubham 2023].
Flow cytometry is central to immune profiling, so researchers need to improve the quality and consistency of flow data. Optimizing study design also helps them draw stronger, more informative conclusions.
Keeping sample collection, handling, and processing consistent can reduce how many study participants you need to find and confirm a biomarker. This matters especially in longitudinal studies with multiple sample collections.
Flow cytometry instruments have gotten more sensitive and accurate, improving data quality and cutting down expert hands-on time. But an often-overlooked source of variability comes from the samples themselves. That’s why researchers and regulatory agencies like the National Institute of Standards and Technology (NIST) are pushing for standardized sample preparation in flow cytometry experiments for therapeutic development.
Quality Samples, Quality Data
High-quality flow cytometry data starts with the source — your data are only as good as the samples you evaluate. Inconsistent sample quality, caused by uncontrolled differences in acquisition, delivery, storage, and processing, can quietly skew the accuracy and variability of your results. That leads to weaker conclusions and limits how much you can translate the findings.
Recent studies show how these inconsistencies create real problems with real consequences for flow cytometry results:
- The method and duration of peripheral blood mononuclear cell (PBMC) cryopreservation — a common step in identifying immune cell clinical biomarkers — changes immune cell population, viability, and reactivity in later flow cytometry analysis. That degrades the accuracy of results [Li 2022, Ticha 2021].
- PBMC processing delays as short as 24 hours can increase granulocyte contamination in flow cytometry analysis — a reminder of why consistent, timely processing after collection matters [Yi 2023].
Sanguine’s onsite prospective collection services let researchers process samples in as little as 2 hours after collection. That’s the value of prospective collection: it avoids processing delays and prolonged PBMC storage.
Despite the clear benefits of prospective collection for flow cytometry, most researchers still source specimens from biobanks, which limits standardization and control over collection and processing.
These indirect approaches are also less convenient for patients and healthy donors. They require donors to give samples at medical facilities instead of at home or work, which limits how geographically and demographically diverse the donor network can be.
Put together, a direct-to-patient strategy benefits translational research by meeting the needs of both the researcher and the patient, leading to more informative and meaningful conclusions.
Longitudinal Study Design for More Informative Immunoprofiling
Longitudinal studies in therapeutic development give you more statistical power from fewer participants, while letting you draw stronger conclusions based on real-world data that resembles later clinical trials.
Longitudinal studies have also proven useful for flow cytometry-based immune profiling in therapeutic development. For example, one recent study used a longitudinal approach to track changes in B and T-cell responses in multiple sclerosis (MS) patients after a booster dose of a COVID-19 mRNA vaccine.
Researchers analyzed whole blood from MS patients and healthy controls by flow cytometry, six months after the primary vaccine and again 4-6 weeks after the booster. They found MS patients had higher levels of terminally differentiated CD4+ and CD8+ effector memory cells than healthy controls [Aiello 2023].
These findings showed the mRNA vaccine works to prevent COVID-19 in MS patients. They also shed light on the potential value of giving these therapies to MS patient populations.
A major challenge in running good longitudinal studies is limited access to recallable donors. Sanguine’s response is a direct-to-patient approach, including onsite and at-home collection services. This makes sample procurement more streamlined and convenient, and makes patient retention and recallability more realistic.
This flexibility also lets researchers engage large networks of patient donors by putting donor needs first, which further strengthens longitudinal studies.
Simultaneous Sample Collection
The immune system is a complex mix of cells and cytokines that no single assay or sample type can fully capture.
Flow cytometry of blood samples and PBMCs is one of the most common approaches to immune profiling. But researchers typically need complementary assays to fully characterize immune profiles for therapeutic development.
For example, in the PBMC processing-delay study above, Yi and colleagues used single-cell RNA sequencing and ELISpot to identify changes in gene expression and interferon-γ secreting cells from stimulated PBMC ex vivo [Yi 2023].
Immunoprofiling analysis often needs multiple sample types beyond PBMC.
A team from Sana Biotechnology published a 2023 study on editing primary pancreatic islet cells (p-islets) from type 1 diabetes mellitus donors (T1DM) to make them better suited for therapeutic engraftment.
Researchers have explored allogeneic engraftment of p-islets to treat T1DM, but these treatments often trigger an immune response from the host, leading to rejection.
In this study, the team engineered donor p-islets to overexpress CD47, making the cells hypoimmune and lowering the risk of rejection after transplant.
To test host immune response and function against the engineered allogeneic p-islets, the team built a preclinical, humanized T1DM mouse model using PBMC and sera from T1DM patients in Sanguine’s donor community [Hu 2023].
Using flow cytometry, the researchers confirmed a higher abundance of CD47+ cells within the engineered p-islets compared to autologous controls after transplantation — a sign the cells kept their hypoimmune properties after engraftment.
Bringing the Patient Experience to Flow Cytometry Assays
To make immune profiling experiments more valuable, Sanguine has built a series of simple but effective solutions that give researchers more confidence in their data and results.
Sanguine addresses unintended sample variability by letting researchers dictate and standardize rapid and flexible sample procurement and processing across sample types, including blood, PBMC, leukopaks, saliva, stool, and urine.
Sanguine also gives researchers unmatched access to patient data and sample procurement from engaged, large, and diverse healthy and disease-state donor populations. This gives researchers the flexibility and capability to run robust prospective and longitudinal immune profiling studies.
Find out more about how Sanguine can help you power more precise and practical flow cytometry and immune profiling assays for your research needs.
By: William Lawrence, Ph.D.; Geocyte
References
[1] Pushparaj, PN. (2020) Chapter 11 – Translational interest of immune profiling. Precision Medicine for Investigators, Practitioners and Providers. 105-122 DOI: https://doi.org/10.1016/B978-0-12-819178-1.00011-3
[2] Shubham, S. (2023). Flow Cytometry Market Research 2032. Allied Market Research Group https://www.alliedmarketresearch.com/flow-cytometry-market
[3] Li, B. (2022) Comprehensive evaluation of the effects of long-term cryopreservation on peripheral blood mononuclear cells using flow cytometry. BMC Immunology DOI: 10.1186/s12865-022-00505-4
[4] Ticha, O. (2021) Effects of long-term cryopreservation of PBMC on recovery of B cell subpopulations. Journal of Immunological Methods. DOI: 10.1016/j.jim.2021.113081
[5] Yi PC. (2023) Impact of delayed PBMC processing on functional and genomic assays, Journal of Immunological Methods. Volume 519, 113514, ISSN 0022-1759, DOI: https://doi.org/10.1016/j.jim.2023.113514
[6] Aiello A. (2023) Longitudinal characterisation of B and T-cell immune responses after the booster dose of COVID-19 mRNA-vaccine in people with multiple sclerosis using different disease-modifying therapies. Journal of Neurology, Neurosurgery & Psychiatry. Volume 94:290-299 DOI: 10.1136/jnnp-2022-330175
[7] Hu X. (2023) Human hypoimmune primary pancreatic islets avoid rejection and autoimmunity and alleviate diabetes in\allogeneic humanized mice. Sci Transl Med. Vol 12;15(691) DOI: 10.1126/scitranslmed.adg5794
