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Adapting the SMART tube technology for flow cytometry in feline full blood samples
Katharina Zwicklbauer1, Dominik von la Roche2, Daniela Krentz1
1LMU Small Animal Clinic, Centre for Clinical Veterinary Medicine, LMU Munich, Munich, Germany.
Insights
A new feline whole blood protocol using SMART Tubes enables long-term storage for flow cytometry analysis. This veterinary research tool allows immune cell quantification up to two years post-sampling.
Area of Science:
- Veterinary immunology
- Clinical pathology
Background:
- Flow cytometry is crucial for monitoring immune responses in humans and cats, particularly for feline immune deficiency virus (FIV).
- Prompt processing of fresh feline blood is often required for flow cytometry, limiting analysis of stored samples from clinical studies.
Purpose of the Study:
- To adapt and validate the SMART Tube (SMT) system for long-term preservation and flow cytometry analysis of feline whole blood.
- To assess the stability and accuracy of immune cell quantification from stored feline blood samples.
Main Methods:
- A novel flow cytometry protocol using SMT for feline whole blood was developed and optimized.
- Quantification of T-helper cells, cytotoxic T-cells, B-cells, monocytes, and neutrophils was performed on samples stored up to two years.
- Results from frozen-stabilized samples were compared with fresh samples and conventional hematology analyzer counts.
Main Results:
- The SMT protocol successfully enabled flow cytometry analysis of feline whole blood samples stored for up to two years.
- Quantification of major immune cell populations (T-helper, cytotoxic T-cells, B-cells, monocytes, neutrophils) was reliable.
- The method was validated against fresh samples and demonstrated clinical applicability in a feline infectious peritonitis (FIP) study.
Conclusions:
- The adapted SMT protocol is suitable for analyzing stored feline whole blood samples via flow cytometry.
- This technique facilitates long-term storage and analysis, opening new research avenues in feline immunology and clinical studies.
Abstract:
Flow cytometry of blood samples is a very valuable clinical and research tool to monitor the immune response in human patients. Furthermore, it has been successfully applied in cats, such as for infections with feline immune deficiency virus (FIV). However, if cells are not isolated and frozen, analysis of anticoagulated blood samples requires mostly prompt processing following blood collection, making later analysis of stored full blood samples obtained in clinical studies often impossible. The SMART Tube system (SMART TUBE Inc., California, United States; SMT) allows fixation and long-term preservation of whole blood samples at -80°C. However, this system has so far only been applied to human biological samples. In the present study, a new flow cytometry SMART Tube protocol adapted for feline whole blood samples was successfully established allowing quantification of T-helper cells, cytotoxic T-cells, B-cells, monocytes, and neutrophils up to 2 years post sampling. Results obtained from frozen stabilized and fresh blood samples were compared for validation purposes and correlated to differential blood counts from a conventional hematology analyzer. Clinical applicability of the new technique was verified by using samples from a treatment study for feline infectious peritonitis (FIP). Using the new SMT protocol on retained samples, it could be demonstrated that long-term storage of these SMT tubes is also possible. In summary, the newly adapted SMT protocol proved suitable for performing flow cytometry analysis on stored feline whole blood samples, thus opening up new avenues for veterinary research on a variety of aspects of clinical interest.
