Easy and high titer production of retroviruses from a producer cell line

June 22, 2015

University of Michigan
Pharmaceutical Sciences
Research Fellow

Overall

Quality of Results

Ease-of-Optimization

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Company:

Takara Clontech

Product Name:

Pantropic Retroviral Expression System

Catalog Number:

631512

Generation of retroviral vectors from a producer cell needs to be reliable for downstream applications for therapeutics development. We used the pantropic retroviral expression system to obtain retroviral particles so as to design new hybrid retroviral vectors. See the following papers for our results using this kit. 1. http://pubs.acs.org/doi/abs/10.1021/mp300561y 2. http://www.sciencedirect.com/science/article/pii/S0168365914004696 3. http://www.sciencedirect.com/science/article/pii/S0168365915002333

Experimental Design and Results Summary

Application

Development of viral and hybrid vectors for gene therapy

Starting Material

None

Protocol Overview

The manufacturer's instructions were used to generate a stably transfected producer cell line. Growth media with serum but without antibiotics were used to continue passaging the cell line (not more than 15 passages). Optimization was needed to efficiently obtain a high titer of virus. Typically, 3-4 days was needed, post seeding at 10% confluency to obtain a good titer after which cells would die

Tips

Optimization for retroviral titer is key. Make sure you can save as many cells from the first passage as possible to obtain a fresher batch post thawing once 15 passages are done.

Results Summary

As shown in the image, optimization was key but generally high (1 x 10^9 - 1 x 10^10 retroviral particles/ml)and usable titer of virus was obtained post 3-4 days of incubation without change in media. The viral quantification was done using the Retro-X Quant kit also available from Clontech (See Catalog No. 631453)

Additional Notes

None

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Summary

The Good

Easy to use. Stably transfected cells are a reliable source of retroviruses

The Bad

Optimization of titer can take a while. Less than 15 passages makes it critical to save early passages for fresh thaw.

The Bottom Line

A very popular and stable source of retroviruses for your transduction and therapeutic formulation needs.

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