Screening of anti-human leukocyte monoclonal antibodies for reactivity with equine leukocytes.

The research team tested a range of anti-human leukocyte monoclonal antibodies to see if they could also react with horse leukocytes, as part of their efforts to improve understandings of equine immunology.

Research Methodology

• 379 different monoclonal antibodies (mAbs) initially submitted to the HLDA8 animal homologues section were tested for cross-reactivity with equine leukocytes.
• The testing process involved two rounds of flow cytometric analysis, both initially in one lab and then with the results corroborated in a further two independent labs.
• The testing procedure looked for similarities between the human and equine staining pattern in the antibodies, as this was one possible indication of cross-reactivity.

Initial Findings

• After the first round of testing, 17 mAbs were flagged as potentially cross-reactive.
• This was narrowed down to 14 mAbs directed against CD2, CD11a, CD18, CD44, CD45, CD49d, CD91, CD163 and CD172, in the second round of testing.
• Other findings included:
  • 1 mAb with weak likely positive reactivity.
  • 12 mAbs with positive staining that likely do not reflect valuable data.
  • 5 mAbs with clear alternate expression pattern different from that of humans.
  • 5 mAbs with a questionable staining pattern.
  • 32 mAbs with weak-positive expression and an alternate staining pattern.
  • 279 mAbs showed negative responses.

Shortcomings and Recommendations for Further Research

• The researchers were unable to test certain types of cells, such as thymocytes or stem cells, because they were unavailable for the screening.
• As only 4% of mAbs submitted reacted positively, there is a need for further testing with a greater variety of anti-human mAbs, as well as developing species-specific anti-CD mAb.

Implications of the Study

• This study highlighted the potential value of a “cross-reactivity” approach to advancing equine immunology.
• The findings also serve as a guide for future research in the field, and underscore the need for continued efforts.