College
College of Arts and Sciences
Mentor Information
Dr. Kenneth Shain
Description
Multiple myeloma is a malignancy of plasma cells for which emerging therapies increasingly target B-cell maturation antigen (BCMA). Many experimental approaches used to investigate therapeutic resistance rely on genetically modified cell populations that must be maintained through antibiotic selection to ensure stable expression of introduced constructs. However, antibiotic sensitivity varies across cell lines and drug types, and improper dosing can compromise experimental reproducibility by allowing unmodified cells to persist or inducing excessive cytotoxic stress. This study aimed to define efficient and practical selection conditions for two commonly used antibiotics, G418 and puromycin, in Jurkat and TALL-104 human T-cell lines. Cells were exposed to serial dilutions of each antibiotic over a 10–11 day period, and viability was assessed using a CCK-8 metabolic assay and trypan blue exclusion. Dose-response kill curves were generated, and IC50 values were estimated to evaluate cytotoxic effects. Jurkat T cells demonstrated strong dose-dependent sensitivity to both antibiotics, with puromycin producing rapid cytotoxicity at low concentrations and G418 exhibiting a more gradual response. In contrast, TALL-104 cells displayed markedly reduced sensitivity, particularly to puromycin, with cytotoxic effects observed only at substantially higher concentrations for G418. Analysis of stabilized time points revealed more consistent trends compared to early variability. These findings demonstrate that antibiotic selection is strongly cell line dependent and emphasize the importance of empirically defining optimized selection ranges to improve reproducibility and support the reliable generation of genetically modified models for multiple myeloma research.
Optimization of Antibiotic Selection Conditions in Human T-Cell Lines to Study Mechanisms of Resistance to Bi-specific Antibody Therapy in Multiple Myeloma
Multiple myeloma is a malignancy of plasma cells for which emerging therapies increasingly target B-cell maturation antigen (BCMA). Many experimental approaches used to investigate therapeutic resistance rely on genetically modified cell populations that must be maintained through antibiotic selection to ensure stable expression of introduced constructs. However, antibiotic sensitivity varies across cell lines and drug types, and improper dosing can compromise experimental reproducibility by allowing unmodified cells to persist or inducing excessive cytotoxic stress. This study aimed to define efficient and practical selection conditions for two commonly used antibiotics, G418 and puromycin, in Jurkat and TALL-104 human T-cell lines. Cells were exposed to serial dilutions of each antibiotic over a 10–11 day period, and viability was assessed using a CCK-8 metabolic assay and trypan blue exclusion. Dose-response kill curves were generated, and IC50 values were estimated to evaluate cytotoxic effects. Jurkat T cells demonstrated strong dose-dependent sensitivity to both antibiotics, with puromycin producing rapid cytotoxicity at low concentrations and G418 exhibiting a more gradual response. In contrast, TALL-104 cells displayed markedly reduced sensitivity, particularly to puromycin, with cytotoxic effects observed only at substantially higher concentrations for G418. Analysis of stabilized time points revealed more consistent trends compared to early variability. These findings demonstrate that antibiotic selection is strongly cell line dependent and emphasize the importance of empirically defining optimized selection ranges to improve reproducibility and support the reliable generation of genetically modified models for multiple myeloma research.
