The research investigates the optimal storage parameters for conducting assays to detect surface-associated immunoglobulin on red blood cells and platelets in horses using flow cytometry. It found that the percentage of red blood cells with surface-associated immunoglobulin remained consistent for up to 72 hours of storage, while for platelets, testing should optimally be conducted in under 4 hours after collection, or be stored at 4°C and tested no later than 24 hours after collection.

Objective

This study sought to determine the optimal storage time and temperature for the detection of surface-associated immunoglobulin (sIg) on red blood cells and platelets in horses using flow cytometry. This is because differentiating between immune-mediated causes and other causes of anemia and thrombocytopenia, a common blood disorder, can be a challenge.

Methodology

• Flow cytometry assays were performed on blood samples from healthy horses at 0, 4, 24, 48, and 72 hours post collection.
• Red blood cell samples were stored at 4°C, while platelet samples were stored at 4°C and room temperature.

Findings

• The study found that the percentage of red blood cells with surface-associated immunoglobulin remained consistent for up to 72 hours of storage.
• However, the percentage of surface-associated IgG on platelets increased above baseline at all time points. The percentage of positive platelets was inconsistent across time points for IgM and IgA.
• Therefore, ideally, platelet testing should be conducted within 4 hours of collection, and in cases where this is not feasible, samples should be stored at 4°C and tested no later than 24 hours after collection.

Conclusion and Recommendations

• The research recommends that while the cutoff values for red blood cells with surface-associated immunoglobulin remained the same across time points, results for platelets should be compared to specific time-based cutoff or reference intervals established by the laboratory conducting the test.
• For more accurate results, laboratories are encouraged to conduct platelet testing ideally within 4 hours of sample collection and if this is not possible, to store samples at 4°C and test no later than 24 hours post collection.