Virus particle purification?

More Deemah Dabbagh's questions See All

Purification of AAV by ion exchange chromatography. How to adjust pH of virus lysate?

I would like to purify AAV particles using ion exchange chromatography. I am testing different pH levels for elution. Is it ok (safe for the virus particles) to use HCL or NaoH to...

08 September 2018 9,089 1 View

AAV purification protocol using chromatorgraphy columns for serotype 8?

Does anyone have a protocol (that has worked) or can suggest a protocol of AAV8 purification using an ion exchange column or any type of chromatography. I am looking for a procedure that does not...

31 December 2017 6,120 1 View

Density gradient- and serotype-independent method of AAV purification?

Dear all, Has anyone tried purifying recombinant AAV virus without density gradient ultracentrifugation in a serotype indepedent manner. Also, Is there a way to achieve high titer without...

11 December 2017 505 4 View

Does any one know if there are anti-HBV antibodies for western blot that recognize multiple HBV antigens?

It would be helpful if the antibody can be used for ELISA and Flow cytometry also.

02 March 2016 1,622 2 View

What is the best/cost-effective/fastest method for determining the titer of AAV?

So, I have transfected 293HEK cells with AAV-GFP vector and after purification, I ended up with 50ul of purified virus (or so I hope). What would be the best method to determine my titer with the...

01 February 2016 5,015 5 View

Can someone help with the tubular structures formed by bunyaviruses during their maturation?

Are these structures part of the Golgi membrane itself? or are they independent viral structures?

08 September 2015 1,308 1 View

Why might my GFP tag be dissociating from my plasmid after transfection?

I have a set of stably transfected cell lines all transfected with plasmids containing GFP tags on the C terminus. During a western blot using anti-GFP antibody, one of my plasmids has dissociated...

01 March 2021 9,310 4 View

Can someone suggest a way to prepare slides to observe under confocal microscope for observing fluorescent tagged proteins??

I am growing the cells on coverslips and transfecting the fluorescent tagged protein directly on them. Then i want to observe these cells under confocal microscope. Currently i am just using...

01 March 2021 6,142 3 View

How vaccines of covid 19 work?

Millions ask how vaccines of covid 19 be successful and safe, its here to explain generally how vaccine of pfizer work ,mRNA vaccines teach our cells how to make a protein that will trigger an...

28 February 2021 2,723 4 View

Are protein samples in SDS compatible with ELISA?

I have used an AllPrep DNA/RNA/Protein Mini Kit QIAGEN kit to extract RNA and protein from my samples. At the end of the protein purification, I resuspend my protein in 5% SDS. Will these samples...

28 February 2021 7,370 3 View

Problem with shRNA transfection. Why are only two plates getting contaminated instead of all?

I transfected my LNCaP-WT cells with 3 shRNA plus their NTC two weeks ago and split two puromycin selected cell plates on Friday last week(Feb 26). I checked for GFP in the cells, and they all...

28 February 2021 4,949 3 View

Does RNAiMAX transfection reagent work in serum free medium?

I diluted siRNA and RNAiMAX in opti-MEM and added to the cells which they were in the growth medium. Is it a right way? or should I culture cells in the opti-MEM medium for a while and not in...

26 February 2021 10,041 3 View

How to replicate the well conditions of a crystal hit with 20%(w/v) isopropanol?

I had a crystal hit in a well with 0.1M HEPEs pH 7.5, 10%(w/v) PEG 4000, and 20%(w/v) isopropanol. Why is isopropanol in w/v here if it's just solvent? How do I recreate this well condition?

25 February 2021 9,761 1 View

How do I transfect a large plasmid into PC12 cells with nerve growth factor (NGF) receptor?

I have a large 16 kb plasmid, which I need to transfect into PC12 cells. Lipofectamin 2000 didn't work and with GFP alone the transfection rate is very low. I also tried the neon invitrogen...

25 February 2021 6,635 5 View

What is a good reagent for lentiviral plasmid transfection in HeLa cells?

I have been using Lipofectamine 2000 but was hoping there may be a better protocol or one for something like Lipofectamine PLUS reagent.

24 February 2021 1,633 4 View

Best plasmid could be used as a standard curve for AAV titration?

I have been trying to have a good AAV titration for my production and I have tried different plasmids to set up as a standard curve but each one behaves differently. i am trying to use ITR for...

23 February 2021 5,167 3 View

Syed A Ali

Hi,

You may want to go through this paper:

http://www.nature.com/articles/mtm201434

Samantha Nicholson

There are several methods for purifying AAV these include cesium chloride or iodixonal gradient, the previously mentioned paper is an excellent place to start you can also have a look at the Balazs paper from 2013

Article Broad protection against influenza infection by vectored imm...

Deemah Dabbagh

Thanks!

Benjamin Strobel

Hi Deemah,

we also just published a comparative method paper on the two most commonly used purification methods, i.e. cesium chloride and iodixanol. You might wanna check this out as well... good luck!

Article Comparative Analysis of Cesium Chloride- and Iodixanol-Based...