Tracking the Total CD8 T Cell Response Following Whole Plasmodium Vaccination

Abstract

CD8 T cells are critical mediators of protection against Plasmodium liver-stage infection. Most studies of the CD8 T cell response to whole parasite Plasmodium vaccines address a single T cell epitope in BALB/c mice, and thus provide limited information. Here, we describe a surrogate activation marker approach that uses the coordinate downregulation of the CD8α chain and upregulation of the integrin CD11a to track the total CD8 T cell response to Plasmodium vaccination via flow cytometry. With this approach, quantitative (magnitude, kinetics) and qualitative (distribution, phenotype, and function) features of the total CD8 T cell response to vaccination with attenuated Plasmodium or other pathogens can be studied.

Trimethoprim-Sulfamethoxazole Prophylaxis During Live Malaria Sporozoite Immunization Induces Long-Lived, Homologous, and Heterologous Protective Immunity Against Sporozoite Challenge

“…Briefly, splenic cells were buffer. Cells were washed twice with Hank's balanced salt solution (HBSS, Sigma Aldrich, St. Louis, Missouri) and suspended in complete medium (Iscove's modified Dulbecco's medium supplemented with 10% fetal bovine serum, glutamine, Pen Strep, and HEPES) [27]. Hepatic lymphocytes were obtained by a method described previously with few modifications.…”

“…The supernatant was then discarded, and the pellet was suspended in 5 mL of ACK lysing buffer for 3 minutes. HBSS was added, and the cells were centrifuged at 400×g for 7 minutes twice, followed by suspension in complete medium [27].…”

“…Specifically, circumsporozoite protein (CS)-specific T cells have been shown as sufficient to confer protection against sporozoite challenge in the irradiated sporozoite model [27]. To identify CD8 + T cells responding to TMP-SMX chemoprophylaxis vaccination, we used the H-2K d -restricted immunodominant P. yoelii CS epitope SYVPSAEQI peptide (derived from P. yoelii CS 280-288 ) [31]) as an indicator by measuring IFN-γ release by responding cells.…”

“…Plasmodium antigens in addition to CS epitopes [32], we used the previously described CD11a hi CD8α phenotype, which has been validated as a surrogate for identification of the global CD8 + T-cell response to whole-organism vaccination [27].…”

“…Vaccine-induced CD8 + T-cell populations were characterized using the following methods, as previously described [27,33]. Cells were first stained with Violet LIVE/DEAD fixable dead cell stain (Invitrogen) to exclude dead cells from the analysis.…”