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Updated: Aug 21, 2026
Preparation of Peripheral Blood Mononuclear Cell Pellets and Plasma from a Single Blood Draw at Clinical Trial Sites for Biomarker Analysis
Published on: March 20, 2021
A simple method to optimize peripheral blood mononuclear cell preparation from cynomolgus monkeys and improve mixed
Marc Bigaud1, Claudine Maurer, Corrine Vedrine
1Novartis Pharma AG, Transplantation Research, WSJ-386 1.55A, CH-4002 Basel, Switzerland. marc.bigaud@pharma.novartis.com
Introduction:
The standard Ficoll-Hypaque method used to isolate peripheral blood mononuclear cells (PBMC) gives very variable results when used with cynomolgus monkey blood. We have improved the method by using special cell processing tubes (CP-tubes), originally developed for clinical use.
Methods:
Blood samples were collected from cynomolgus monkeys and processed for PBMC preparation using either the classical Ficoll-Hypaque method or CP-tubes following various centrifugation protocols. The preparations were compared according to their cellular content as well as their response in the mixed lymphocyte reaction (MLR).
Results:
Good PBMC separation was achieved in >90% of samples by centrifugation of blood samples in CP-tubes for 40-45 min at 1650 x g and 20 degrees C. For the remaining samples, poor PBMC separation was probably due to low corpuscular hemoglobin concentrations (< 28 g/dl), but this could be rectified by using one to two additional centrifugations. The PBMC preparations thus obtained showed lower red blood cell (RBC) and polymorphonuclear (PMN) cell contamination, reacted well to mitogen and showed improved MLR stimulation indices vs. standard Ficoll-Hypaque-PBMC-derived preparations. The inhibitory effect of Cyclosporine-A (CsA) was within the low nanomolar range with both methods.
Discussion:
These results demonstrate that the use of CP-tubes is a practical way of improving PBMC separation and MLR responses with cynomolgus monkey blood.
Insights
Improved cell processing tubes (CP-tubes) enhance peripheral blood mononuclear cell (PBMC) isolation from cynomolgus monkey blood, yielding better mixed lymphocyte reaction (MLR) responses compared to the standard Ficoll-Hypaque method.
Area of Science:
- Immunology
- Primate Research
- Cell Biology
Background:
- The standard Ficoll-Hypaque method for isolating peripheral blood mononuclear cells (PBMC) yields inconsistent results with cynomolgus monkey blood.
- A novel approach using specialized cell processing tubes (CP-tubes) was investigated to improve PBMC isolation efficiency.
Purpose of the Study:
- To optimize and validate a modified Ficoll-Hypaque method using CP-tubes for reliable PBMC isolation from cynomolgus monkey blood.
- To compare the quality of PBMC preparations and their functional responses (Mixed Lymphocyte Reaction - MLR) obtained by the standard and modified methods.
Main Methods:
- Cynomolgus monkey blood samples were processed for PBMC isolation using both the classical Ficoll-Hypaque technique and CP-tubes with varied centrifugation conditions.
- PBMC preparations were evaluated based on cellular yield, purity (contamination by red blood cells and polymorphonuclear cells), and their proliferative response in MLR assays.
Main Results:
- CP-tubes achieved successful PBMC separation in over 90% of samples with optimized centrifugation parameters (40-45 min, 1650 x g, 20°C).
- PBMC isolated using CP-tubes exhibited lower red blood cell and polymorphonuclear cell contamination.
- These preparations demonstrated robust responses to mitogens and significantly improved MLR stimulation indices compared to standard Ficoll-Hypaque method.
Conclusions:
- The use of CP-tubes offers a practical and effective method for enhancing PBMC isolation from cynomolgus monkey blood.
- This improved method leads to superior PBMC quality and more reliable functional responses in MLR assays, crucial for preclinical research.
