Anti-M. tuberculosis Proteinderivative Monoclonal antibody, Clone CDI568

To evaluate the negative regulatory effect of human CD4(+)CD25(+) regulatory T cells (Treg) on anti-Tuberculosis (TB) immune responses of T cells including CD4(+), CD8(+) and V gamma 2V delta 2(+) T cells simultaneously in vitro, a versatile carboxyfluorescein succinimidyl ester (CFSE)-based proliferation assay was exploited to determine the suppressive effect of Tregs on the purified protein derivative (PPD)-specific proliferations of such T-subsets in peripheral blood mononuclear cells (PBMC) of BCG-infected donors. We demonstrated that PPD-stimulation droved synchronously the proliferations of both CD4(+) and CD8(+) T cells and V gamma 2V delta(+) T cells in PBMCs. Such PPD-specific proliferations of CD4(+), CD8(+) and V gamma 2V delta(+) T cells can be prohibited by human Treg concurrently. Similarly, Treg has the ability of suppressing the CD3/CD28 antibody-induced proliferations of such T-subsets. Our results indicated that the versatile CFSE-based proliferation assay can be applied to determine the complicated anti-TB immune response of multi-T-subsets simultaneously, and may facilitate human Tregs as potential tool to down-regulate overdue anti-TB immune responses involved in multiple effective T-subsets to enhance protection against Mycobacterium tb (M. tb) infection. [Qin J, Gong GM, Sun SL, Song B, Du Y, Yang X, Wang N, Zhu S, Xuan XY, Liu PP, Xu YM. Simultaneous determination of Human CD4(+)CD25(+) regulatory T cells suppressing anti-TB immune responses of CD4(+), CD8(+) and V gamma 2V delta(+) T cells in vitro. Life Sci J 2012; 9(3):551-555] (ISSN: 1097-8135). http://www.lifesciencesite.com. 77

The need for an accurate, rapid, simple and affordable point-of-care (POC) test for Tuberculosis (TB) that can be implemented in microscopy centers and other peripheral health-care settings in the TB-endemic countries remains unmet. This manuscript describes preliminary results of a new prototype rapid lateral flow TB test based on detection of antibodies to immunodominant epitopes (peptides) derived from carefully selected, highly immunogenic M. tuberculosis cell-wall proteins. Peptide selection was initially based on recognition by antibodies in sera from TB patients but not in PPD-/PPD+/BCG-vaccinated individuals from TB-endemic settings. The peptides were conjugated to BSA; the purified peptide-BSA conjugates striped onto nitrocellulose membrane and adsorbed onto colloidal gold particles to devise the prototype test, and evaluated for reactivity with sera from 3 PPD-, 29 PPD+, 15 PPD-unknown healthy subjects, 10 patients with non-TB lung disease and 124 smear-positive TB patients. The assay parameters were adjusted to determine positive/negative status within 15 minutes via visual or instrumented assessment. There was minimal or no reactivity of sera from non-TB subjects with the striped BSA-peptides demonstrating the lack of anti-peptide antibodies in subjects with latent TB and/or BCG vaccination. Sera from most TB patients demonstrated reactivity with one or more peptides. The sensitivity of antibody detection ranged from 28-85% with the 9 BSA-peptides. Three peptides were further evaluated with sera from 400 subjects, including additional PPD-/PPD+/PPD-unknown healthy contacts, close hospital contacts and household contacts of untreated TB patients, patients with non-TB lung disease, and HIV+TB- patients. Combination of the 3 peptides provided sensitivity and specificity >90%. While the final fully optimized lateral flow POC test for TB is under development, these preliminary results demonstrate that an antibody-detection based rapid POC lateral flow test based on select combinations of immunodominant M. tb-specific epitopes may potentially replace microscopy for TB diagnosis in TB-endemic settings.