Human leukemia T-cell lines as alternative to animal use for detecting biologically active Staphylococcal enterotoxin type B

Authors: Rasooly, Reuven; Do, Paula; He, Xiaohua; Hernlem, Bradley - Brad

Submitted to: Toxins
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/21/2021
Publication Date: 4/23/2021
Citation: Rasooly, R., Do, P.M., He, X., Hernlem, B.J. 2021. Human leukemia T-cell lines as alternative to animal use for detecting biologically active Staphylococcal enterotoxin type B. Toxins. 13(5). Article 300. https://doi.org/10.3390/toxins13050300.
DOI: https://doi.org/10.3390/toxins13050300

Interpretive Summary: Food poisoning is the result of food becoming contaminated with toxins that make people sick. A common source of such toxins is the Staphylococcal bacteria which can make a large number of different but related toxins. There are many good ways to tell whether a toxin is present in food but to tell whether it is in a form that can make people sick usually involves live animal testing which is ethically disfavored. We report a method to detect and measure the amount of a particular toxin, Staphylococcal enterotoxin type B (SEB), using human cells that can be kept alive in the lab instead of using or killing live animals. We show that this method can measure very low amounts of SEB and that the method responds particularly to SEB and not to other similar toxins. The importance of this work is in food safety as a way to tell whether SEB is present and in a form that can make people sick as well as a way to tell if food processing, such as heat pasteurization, has been effective to make any toxins present inactive.

Technical Abstract: Staphylococcal enterotoxin type B (SEB) is associated with food poisoning. The present detection methods for biologically active SEB rely on the emetic response in live kittens or monkeys. This technique suffers from poor reproducibility, low sensitivity and causes ethical concerns regarding the use laboratory animals. The data presented here show, for the first time, the successful alternative to live animal testing that utilizes SEB superantigenic activity inducing cytokine production for specific novel cell-based assay for quantifiable detection of active SEB. Rather than using or sacrificing live animals, we found that SEB bind to major histocompatibility complex (MHC) class II molecules on Raji B-cells and present this SEB-MHC class II complex to the specific V-ß5.3 regions of human T-cell line HPB-ALL led to a dose dependent secretion of IL-2, capable of being quantified and can detect 10 pg/ml of SEB. This new assay is 100,000 times more sensitive than the ex vivo murine splenocyte method that has achieved the detection limit of 1 µg/ml. The data presented here also demonstrated that SEB induced proliferation in a dose dependent manner of cells obtained by three various selection methods: splenocyte cells containing 22% CD4+ T cell, on CD4+ T cells enrich to > 90% purity by negative selection methods and on CD4+ T cell enriched to > 95% purity by positive selection methods. The highly enriched positively isolated CD4+ T with the lowest concentration of APC cells (below 5%) provided higher cell proliferation than splenocyte cells containing the highest concentration of APC cell.