Degranulation Response in Cytotoxic Rat Lymphocytes Measured with a Novel CD107a Antibody

Abstract

Measuring degranulation through CD107a expression has become an advantageous tool for testing the functional capacity of cytotoxic cells. Such functional studies have been hampered in the rat by the lack of a suitable anti-rat CD107a antibody. In this study, we report a novel hybridoma generated by immunizing Armenian inbred hamsters with transfected Chinese hamster ovary cells expressing CD107a. The SIM1 clone exhibited specific reactivity with CD107a and measured degranulation from natural killer (NK) cells stimulated with target cells or mAb crosslinking of their activating receptors. Degranulation in IL-2-activated NK cells could also be measured, when using low effector to target ratios. SIM1 also stained activated CD8, but not CD4 T cells. This report characterizes the degranulation response in cytotoxic rat cells with a new antibody against rat CD107a.

Signature quality attributes of CD146+ mesenchymal stem/stromal cells correlate with high therapeutic and secretory potency

“…CD107a/LAMP1 (lysosome associated membrane protein-1) expression and functional implications on BM-MSC are largely unknown, until now. CD107a expressed intracellularly on endosomal membranes have been reported on immune cells, most notably cytotoxic T cell and natural killer cells, as indicators of activation or degranulation, respectively (Alter et al, 2004;Kannan et al, 1996;Sudworth et al, 2016;Vego et al, 2016;Wattrang et al, 2015;York and Milush, 2015). Likewise, our data suggests a similar CD107a expression correlative with an activated and secretory function after inflammatory challenge.…”

“…Additional signature markers we describe herein as correlative with CD146 + BM-MSC are CD107a, CXCR4, and LepR. CD107a, or lysosomal-associated membrane protein-1 (LAMP-1), has been described as a marker of highly secretory cytotoxic T cells and actively degranulating natural killer (NK) cells (Alter et al, 2004;Kannan et al, 1996;Sudworth et al, 2016;Vego et al, 2016;Wattrang et al, 2015;York and Milush, 2015), but largely unknown in MSC. Thus, an assessment of the secretory status of the cells (e.g., paracrine activity) would inform about how responsive they are at a particular time, susceptible to be associated with their potency.…”

Gamma/Delta T Cells in the Course of Healthy Human Pregnancy: Cytotoxic Potential and the Tendency of CD8 Expression Make CD56+ γδT Cells a Unique Lymphocyte Subset

“…We performed this well-established assay (42)(43)(44)(45) as described by Andzelm et al and previously presented in our publications (15,46,47): Cells were incubated for 4 h at 37°C and 5% CO 2 with the fluorochrome-conjugated anti-CD107a antibodies in RPMI 1640, supplemented with 10% FCS, penicillin, streptomycin, ionomycin (1 µg/ml) (Sigma-Aldrich) and phorbol myristate acetate (25 ng/ml) (Sigma-Aldrich). Before labeling with the other monoclonal antibodies the cells were washed and resuspended in RPMI 1640 with 5% FCS.…”

IL-12, IL-15, and IL-18 pre-activated NK cells target resistant T cell acute lymphoblastic leukemia and delay leukemia development in vivo

“…With this antibody 10-15% degranulation is measured against potent target cells. 55 Degranulation by human NK cells were measured by incubating 100 mL PBMC (2 £ 10 6 cells/mL) with 100 mL K562 target cells (2 £ 10 6 cells/ mL) in the presence of anti-human CD107a-BV510 (BD Biosciences, cat.no. 563078) for 6 h, with Brefeldin A (Sigma-Aldrich, B7651) added at 5 mg/mL for the last 5 h. NK cell cytotoxicity was measured with a standard 51 Cr release assay, using enriched NK cells as effector cells and 51 Cr-labeled YAC-1, YB2/0, or RL as target cells as described previously.…”