Measurement of in vitro leucocyte mitogenesis in fish: ELISA based detection of the thymidine analogue 5′-bromo-2′-deoxyuridine

David T. Gauthier, Deborah D. Cartwright, Christine L. Densmore, Vicki Suzette Blazer, Christopher A. Ottinger

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

In this study we present a method for the measurement of in vitro mitogenesis in fish leucocytes that is based on the incorporation of the thymidine analogue 5′-bromo-2′-deoxyuridine (BrdU) into the DNA of replicating cells, followed by ELISA-based detection. This technique, adapted from methods developed for mammalian cells, operates on a similar biological principle to 3H-thymidine incorporation, but circumvents the logistical and safety issues inherent with the radioactive label. Because it directly measures DNA proliferation, the assay has advantages over other colorimetric methods that may be strongly influenced by leucocyte metabolic status. Using BrdU incorporation followed by ELISA, we evaluate the responsiveness of rainbow trout (Oncorhynchus mykiss [Walbaum]) leucocytes to the mammalian T-cell mitogen Concanavalin A (Con A) as well as the differential response of white perch (Morone americana [Gmelin]) leucocytes to Con A and pokeweed mitogen. Specific considerations intrinsic to the assay system are discussed, including the implications of utilising enzyme-based detection.

Original languageEnglish (US)
Pages (from-to)279-288
Number of pages10
JournalFish and Shellfish Immunology
Volume14
Issue number4
DOIs
StatePublished - Jan 1 2003

Fingerprint

Bromodeoxyuridine
Concanavalin A
thymidine
Thymidine
Fish
Assays
leukocytes
Cells
enzyme-linked immunosorbent assay
Morone americana
Pokeweed Mitogens
T-cells
DNA
concanavalin A
fish
Mitogens
assay
Labels
Oncorhynchus mykiss
Phytolacca americana

All Science Journal Classification (ASJC) codes

  • Environmental Chemistry
  • Aquatic Science

Cite this

Gauthier, David T. ; Cartwright, Deborah D. ; Densmore, Christine L. ; Blazer, Vicki Suzette ; Ottinger, Christopher A. / Measurement of in vitro leucocyte mitogenesis in fish : ELISA based detection of the thymidine analogue 5′-bromo-2′-deoxyuridine. In: Fish and Shellfish Immunology. 2003 ; Vol. 14, No. 4. pp. 279-288.
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Measurement of in vitro leucocyte mitogenesis in fish : ELISA based detection of the thymidine analogue 5′-bromo-2′-deoxyuridine. / Gauthier, David T.; Cartwright, Deborah D.; Densmore, Christine L.; Blazer, Vicki Suzette; Ottinger, Christopher A.

In: Fish and Shellfish Immunology, Vol. 14, No. 4, 01.01.2003, p. 279-288.

Research output: Contribution to journalArticle

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