(Fwd) RE: T cell proliferation kits

[KARIM MAGHNI]

------- Forwarded Message Follows -------
From:          Self <MEAKINS/KARIM>
To:            kevin-ault at uiowa.edu
Subject:       T cell proliferation kits
Date:          Tue, 6 Apr 1999 17:18:11 EST5EDT

Hi Kevin,

We had exactly the same question approximately two years ago. We 
have tested different kits for assessing CD4+ T cell proliferation, 
and then, we have decided to publish our results.
Good luck.

Karim Maghni, Ph.D.

Maghni K, Nicolescu OM, Martin JG. Suitability of cell metabolic colorimetric assays for assessment of
CD4+ T cell proliferation: comparison to 5-bromo-2-deoxyuridine (BrdU)
ELISA. J Immunol Methods 1999 Mar 4;223(2):185-94 .

The conventional tritiated thymidine (3H-TdR) incorporation assay is
considered as the 'gold standard' for the assessment of cell growth.
However, the 3H-TdR incorporation assay has several disadvantages
which have prompted the development of nonradiolabelling proliferation
assays such as 5-bromo-2-deoxyuridine (BrdU) ELISA, tetrazolium
microplate assay and acid phosphatase assay. In studies, these three
proliferation assays have shown equivalent sensitivity and
reproducibility to the 3H-TdR incorporation assay. However the results
may be affected by the cell type studied. In the present study, we
have used these three proliferation assays for the assessment of rat
lymph node CD4 + T lymphocyte growth in response to polyclonal antigen
stimulation. The proliferation assays were compared on the basis of
four criteria: sensitivity, reproducibility, stimulation index and
insensitivity of the assay to the cell number. Our results indicated
that the BrdU ELISA demonstrated the highest sensitivity,
reproducibility and stimulation index but had a limited linear range
for cellular growth. The tetrazolium microplate assay also had a
relatively good sensitivity, reproducibility, stimulation index and a
wider linear response range for cell growth in comparison to the BrdU
ELISA. The acid phosphatase assay showed the lowest reproducibility
and stimulation index. Because BrdU incorporation in DNA of
proliferating cells has been reported to block cell division, we have
investigated this possibility in sequential assays. Our results
indicated that in our experimental conditions no evidence of an
anti-mitogenic action of BrdU was observed. We also compared the
performance of the MTS assay and BrdU ELISA in measuring substance
P-induced CD4 + T cell proliferation. The results indicated that the
MTS assay may reflect change in cell activation leading to an
overestimate of cell growth. In conclusion, our results indicate that
the BrdU ELISA is the most sensitive of the three proliferation assays
used for the assessment of CD4 + T lymphocyte growth and is the
preferred assay when small changes in cell growth are expected.