Page 54 - GPD-2-2

Gene & Protein in Disease

ORIGINAL RESEARCH ARTICLE
Optimization of transfection methods for human

lymphoblast TK6 cell line

Akamu Jude Ewunkem* and Kyle Agee
Department of Biological Science, Winston Salem State University, Winston Salem, North Carolina, USA

Abstract

Transfection has recently gained attention in the field of biomedical research due
to its ability to manipulate gene expression. Every mammalian cell type has a
characteristic set of requirements for optimal transfection. Some cells can be difficult
to transfect and require optimization for successful transfection. Human lymphoblast
TK6 cell line, an important cell line for genotoxic studies, is known to be extremely
hard to transfect. Thus, optimizing transfection methods for human lymphoblast
TK6 are increasingly important. To accomplish this, TK6 human lymphoblasts were
transfected with plasmid constructs that expressed green fluorescent protein (GFP)
and NanoLuc activity. We compared the transfection efficiency of three commercially
®
available transfection reagents, including Amaxa 96-well Nucleofection procedure
using various solutions (SF, SE, and SG), Lipofectamine LTX, and Metafectene Pro . The
®
transfection efficiency and toxicity of various reagents were tested by fluorescence
microscopy, luciferase activity, and cell viability assays. Amaxa 96-well Nucleofection
Solution SF was identified as the best transfection reagent due to its relatively high
luciferase activity, acceptable cell viability (80%), and GFP transfection efficiency
(80%). Optimal conditions for transfection utilized with this reagent included 0.4 µg
of plasmid DNA, 1.8 × 10 cells, and using the DS 137 Nucleofector program.
5
*Corresponding author:
Akamu Jude Ewunkem
(ewunkemaj@wssu.edu) Keywords: Optimization; Nucleofection; Lipofectamine LTX; Metafectene Pro;
Citation: Ewunkem AJ, Agee K, Transfection; Cell viability
2023, Optimization of transfection
methods for human lymphoblast
TK6 cell line. Gene Protein Dis,
2(2):0353.
https://doi.org/10.36922/gpd.0353 1. Introduction
Received: March 21, 2023 Transfection is an important analytical tool for studying the function of genes and
Accepted: May 5, 2023 proteins in a cellular environment [1,2] . Successful transfection is influenced by the quality
Published Online: May 29, 2023 of the nucleic acid, duration of transfection, transfection reagent, and cell lines [3,4] . Many
Copyright: © 2023 Author(s). cell lines can be easily transfected. However, others, such as some human primary cells,
This is an Open Access article are traditionally proven to be very difficult to transfect [5-7] . Cells considered difficult or
distributed under the terms of the
Creative Commons Attribution hard to transfect include, inter alia, stem cells, primary cells, mast cells, natural killer
License, permitting distribution, cells, macrophages, and TK6 human lymphoblasts [8-11] .
and reproduction in any medium,
provided the original work is Successful transfection of thymidine kinase heterozygous cell line of human
properly cited. lymphoblasts (TK6 human lymphoblasts) is a crucial step for studying cell biology at
Publisher’s Note: AccScience the molecular level through gene expression. Many transfection methods have been
Publishing remains neutral with developed. Each transfection method utilizes different approaches. However, each
regard to jurisdictional claims in [12]
published maps and institutional transfection method has advantages and limitations . Viral methods use modified
affiliations. viruses to deliver nucleic acid, while non-viral method uses nanocarriers for delivery. Viral

Volume 2 Issue 2 (2023) 1 https://doi.org/10.36922/gpd.0353