Lipofectamine 3000 Reagent from Thermo Fisher Scientific

January 31, 2017

The Pennsylvania State University
Biochemistry and Molecular Biology
Graduate Student

Overall

Quality of Results

Ease-of-Optimization

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Company:

Thermo Fisher Scientific

Product Name:

Lipofectamine 3000 Reagent

Catalog Number:

L3000015

In our lab, we use Lipofectmamine 3000 reagent as our primary transfection reagent for a wide range of experiments. I, personally, have used this reagent to transfect cells for fluorescent microscopy, GST pull downs, and co-immunoprecipitation experiments. We have had success using Lipofectamine reagents in the past and the Lipofectamine 3000 appears to be the best option.

Experimental Design and Results Summary

Application

Fluorescent Microscopy

Starting Material

MDCK cells

Protocol Overview

I first plated 2 x 10^5 MDCK cells on fibronectin-coated coverslips in a 6-well plate. The next morning with fluorescent constructs using the Lipofectamine 3000 reagent. After 12 hours, I switched the cells to serum-free media. The next morning, I fixed the cells using PFA and mounted them using Prolong Gold. Slides were imaged by deconvolution microscopy.

Tips

In the Lipofectamine 3000 protocol, you can use either 7.5 ul or 5 ul of reagent. I found that 7.5 ul of reagent appears to work the best. I also like to use the higher limit in terms of recommended cell count.

Results Summary

Overall, I really enjoy using Lipofectamine reagent for cell transfections. The protocol is straight forward, easy to use, and does not take up much time. Results are very reproducible when performing multiple experiments.

DOI or PMID #

27562622

Additional Notes

N/A

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Summary

The Good

Very easy to use, fast, good transfection efficiency.

The Bad

Some constructs are more difficult to transfect than others. Some trouble shooting in terms of DNA amounts may need to be performed.

The Bottom Line

Definitely worth trying if you do a lot of transfections in your lab. Easy to learn and easy to use.

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