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Lipo3000 Transfection Reagent

 

Lipo3000 is a type of transfection reagent which is lipid-based in nature and used to transfect cells by transporting the complex molecules into the cell. Due to its cationic nature, it is used in a wide variety of techniques i.e. gene expression or gene delivery.

Gene delivery is a type of procedure in which RNA/DNA is transfected into the cell. For this transfection, lipid-based molecules or cationic liposomes are convenient and efficient for monitoring this RNA or DNA in either somatic cells or germ cells. Generally, for the entry of these cationic liposomes into the cell, they must interact with the negatively charged cell membrane and nucleic acid to suppress these electrostatic forces(Dalby et al., 2004, Douzandegan et al., 2017). Thus after the interaction, the complex of liposome and nucleic acid is formed which provides the entry of a particular gene of interest ( DNA/RNA ) via the electrostatic interaction with the host cell membrane. The Advantage to use cationic liposomes for gene transfer is because of their high efficiency for transfection(Kaestner et al., 2015).

It can transfect a wide variety of cells but it may cause apoptotic effects for some lines of eukaryotic cells. So for the best transfection results, several factors must be under consideration for successful transfection. These include; Plasmid size, target cell type & its number(Douzandegan et al., 2017).

The efficiency of Lipo3000 for KYSE-30 cell line

This KYSE-30 is the cell line of human squamous cell carcinoma. Studies suggest that lipo3000 can be used for transfection of KYSE-30 cell lines with pCDH- 513b vector. It is suggested that the transfection efficiency for the KYSE-30 cell line is improved when lipo3000 is used in a smaller amount. Figure 1 also shows the efficiency of lipo3000 for KYSE-30 cell lines. This flow cytometry data reveals that lipo3000 transfection efficiency is increased when it is used in 1 microliter for 1.5 micrograms of plasmid DNA in KYSE-30 cells which in turn causes a greater level of cellular death in KYSE-30 cell lines. So, the level of plasmid DNA and the level of reagent of transfection play a considerable role in the transfection efficiency and viability of cell lines. It is also reported that during the   transfection in the KYSE-30 cell lines (which are cancerous cells), the increased level of plasmid DNA along with the low level of transfection reagent is required for optimal transfection efficiency.

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Figure 1:The transfection efficiency analysisofpCDH-513b-transfected KYSE-30 cells using 1.5 µg of plasmid DNA. The Flow cytometry was used to analyze the GFP expression, (A) gating for living cells in untransfected cells and (B) untransfected cells for GFP expression.

 It is the novel cationic reagent for transfection, consisting of two main components;

  • LFN 3000
  • P3000

Due to this reason, it is an efficient transfection reagent for entry into the various types of cells.  According to various studies, it is also an efficient transfection reagent to transfer nucleic acids into many types of eukaryotic cells. The cells which are transfected with high transfection efficiency include; fibroblast cells, embryonic stem cells (ESCs), primary cell cultures, and pluripotent stem cells (iPSCs)(Yu et al., 2016).

It can also be used to transfect various types of cargo includes;  siRNA, plasmid DNA, miRNA, etc.(Tamm et al., 2016). 24. Along with these molecules, it is also used to transfect various types of proteins that are exogenous. For example, lipo3000 reagent is used to transfect enzymes which are translation inhibitors into the cancer cells to examine the intracellular processes(Rust et al., 2015, Mahmoudian et al., 2020).

Transfection of mesenchymal stem cells

Although it is a very difficult procedure to transfect the MSCs. Mesenchymal stem cells are multipotent stem cells that are used in different cellular processes. They play an important role to immune protecting the tissues which are injured and can regenerate them via immunomodulatory molecules. The major problem arises when they are used in genetic engineering for an efficient gene delivery process. Different techniques which are used include electroporation, viral-based gene delivery, and non-viral-based gene delivery. Non-viral-based gene transfer includes the use of cationic liposomes. Hence lipo3000 can be used as a transfection reagent for the transfer of molecules into the MSCs. It has almost < 15% gene delivery efficiency that’s why they are considered as difficult to transfect(de Carvalho et al., 2018).

Transfection of HEKC-293 cells:

Different studies suggest that the efficiency of transfection varies as the cell type varies. Depending upon the different conditions, lipo3000 can transfect human embryonic kidney cells HEKC-293 with 10-40% transfection efficiency. Also, it is toxic to HEKC  cells(Rao et al., 2015).

Transfection of HCT-116 cells:

HCT-116 cells are the cell lines of human colon cancer cells. Different studies reported that lipo3000 transfects these cells with almost 50% transfection efficiency(Liu et al., 2017, de Carvalho et al., 2018).

Transfection of U87MG cells:

U87MG cells are the brain cancer cells. They can transfect via the wide Variety of transfection reagents which lipo3000 and lipofection are more suggested ones. Lipo3000 has a 58.21% transfection efficiency for U87MG Cells. On the other hand, lipofectin has 38.64% transfection efficiency with 80% cell viability for brain cancer cells.

Transfection of primary muscle myoblast:

Different transfection reagents can be used for the transfection of primary myoblast cells. These include lipo3000 with a transfection efficiency of 97.78%, lipo2000 with a transfection efficiency of 60.01%, and Fugene with a 77.86% transfection efficiency. So due to the greater efficiency rate, lipo3000 is suggested for transfection of primary muscle myoblast cells.

Transfection of HL-60 cells:

It is a type of Promyelocytic Leukemia Cell line. It shows the restrictions on almost all the transfection reagents. Only the lipo3000 with 8.93% transfection efficiency, and lipo2000 with 7.89% transfection efficiency, can transfect the HL-60 cells. They both have a low level of transfection along with greater cytotoxicity levels i.e. Lipo3000 (51.89% cell viability) & lipo2000 (46.21% cell viability).

Transfection of JU77 cells:

 These are the mesothelioma cell lines of the lungs.  Almost all the transfection reagents can transfect these JU77 cells easily. Between different types of transfection reagents, lipo3000 can transfect the JU77 cells at almost 100% with cellular viability of 68.21%. lipo2000 can also transfect these cells at a higher transfection efficiency of almost 90.21% along with high cell viability of 69.27%.

It is also suggested that the transfection efficiency of all the transfection reagents has a positive correlation with cell toxicity (cytotoxicity).  A transfection reagent with a high-efficiency rate also has a higher level of cytotoxicity(Wang et al., 2018).



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