Brad Huffington

6 Questions 8 Answers 0 Followers

Questions related from Brad Huffington

Can anyone recommend a serum-replacement for mouse embryonic fibroblasts?

We need to culture mouse embryonic fibroblasts without FBS for several days (we want to deprive the cells of serum-derived lipoproteins). The MEFs don't necessarily need to proliferate during this...

04 April 2016 3,438 3 View

Does qRT-PCR amplify ONLY from fully spliced mRNA? Or could it amplify some pre-mRNA that still contains introns?

Does qRT-PCR amplify ONLY from fully spliced mRNA? Or could it amplify some pre-mRNA that still contains introns? I’m not a qRT-PCR expert, and before I reproduce a method from a paper, I want to...

09 September 2015 183 10 View

Western blotting (with ECL reagent and CCD imager): What is the optimum exposure time for bands for quantification?

I'm quantifying proteins on western blots, using chemiluminescent ECL primer and a CCD-based imager (Carestream) to visualize the bands. I can use exposure times of seconds to minutes. The shorter...

04 April 2014 9,390 2 View

How long can you store SDS-PAGE gels before use?

Sometimes I prepare a standard SDS-PAGE gel for westerns, wrap with a wet paper towel and cling wrap, and store it in a zip lock baggy in the fridge. Does storage for a few days alter the...

03 March 2013 9,158 33 View

How to get rid of high background + non-specific negative ("ghost") bands on a western blot of a tissue homogenate?

The antibody (and exact protocol) that I'm using worked in the recent past, but now I get very high background, along with a "ladder" of negative ghost bands against the dark background. My...

01 January 2013 6,801 14 View

Very high molecular weight activity bands in my gelatin zymography?

I'm performing gelatin zymography on my various heart homogenates and extracts. Unfortunately, I'm getting only very faint, or very diffuse, bands at the expected molecular weight. Instead, I'm...

06 June 2012 543 2 View