Identification of koala T lymphocytes using an anti-human CD3 antibody
Wilkinson, R, Barton, M & Kotlarski, I 1995, Developmental and Comparative Immunology, vol. 19, no. 6, pp. 537-545.
Studying the immunobiology of koalas is important because koalas are prone to disease. Recently a new antibody, anti-CD3, has become available. To date koala T cells have not been identified using classic techniques. This study used anti-CD3 antibody to successfully bind T lymphocytes from koala’s peripheral blood. Anti-CD3 was found to bind a small polypeptide, likely similar to the original target – epsilon chain of the human CD3 complex.
Tissues were collected from healthy koalas in captivity. Three tissues were used in the study: blood from alive koalas, formalin preserved tissues such as spleen, kidney and brain from deceased koalas, and cryopreserved or frozen tissue. Immunocytochemistry is a technique that allows labelling tissues with a dye which can then be visualised using a microscope. This technique showed that anti-CD3 antibody bound T and B cells of spleen and lymph nodes in deceased and cryopreserved tissues, but not other tissues such as liver and kidney. Cytometry is used to separate and count different types of cells based on the tag placed on them. This technique identified that once koala blood is fixed, 65% of T and B cells bind anti-CD3 antibody, which is within standard range. Immunoprecipitation and Western Blotting allow isolation of proteins and their binding to specific antibodies to which they would normally bind in the body. These techniques identified a small peptide, 23kDa in size, that bound anti-CD3 antibody. A possible sequence of the peptide was obtained.
The study may be the first in identifying koala T cells from lymphoid organs and blood and in using flow cytometry to enumerate lymphocytes from blood with anti-CD3 antibody. CD3 is a receptor with many peptide chains (such as epsilon) that vary in their similarity between animals. Human epsilon chain is 19kDa while the one identified in the study is 23kDa, suggesting koala CD3 epsilon may be larger and slightly different from the human chain. Better sequencing results are needed to determine the exact sequence of the fragment.
Understanding the biology of the koala’s immune system is important for combating disease. The methodology outlined in the paper can be applied to koala blood, formalin treated and frozen tissues of deceased koalas. Notably, cryopreserved tissue can be used for T cell detection especially if fresh tissue is not available.
Summarised by Alexandra Selivanova
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