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Member since: Jul 28, 2005
From: UK, Scotland
Status: Virology Moderator
My points: 1013    what's this
Name: Jason King

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Jul 1, 2009    11:24 AM

Replied to topic in Stem Cell Biology forum   Commercialisation of AFP

Maybe it would be best to try to commercialize something in an area of biomedical science that you not only understand but are also an expert in.

Jun 29, 2009    12:06 PM

Posted new topic in Cytology, Histology and IHC forum   IF Ab for murine NK cells

Hi, I need to check some cryosectioned tissue for presence of murine NK cells. CD56 seems to be reasonable specific for NK cells. Does anyone have any experience in mouse NK detection in cryosections? ie. Which antibody to choose? There are quite a few clones and most appear not to have been tested ...

Jun 28, 2009    04:55 PM

Replied to topic in DNA (PCR, Real Time PCR) forum   Follow the rules! Get the results!!

This is a useful list - especially rule number 1. How many people really do take 3 (really) independent cell samples for RT-PCR analysis?

Jun 28, 2009    04:50 PM

Replied to topic in Cell Culture and Tissue Culture forum   Thawed cells and subculturing

I thaw my cells (usually about 1mL) and transfer them in to a Falcon tube containing 10mL of fresh medium. I spin the cells down for 3 mins at about 250g (quite gently) then pipette off about 90% of the supernatant. Then add fresh medium again and transfer this and the cells to a TC flask. The day ...

Jun 28, 2009    05:36 AM

Replied to topic in Protein Detection (Western blots, gels, IP) forum   SDS page problem

The most important thing is to ensure that your plates are clean. I use a weak SDS solution, water and then 70% ethanol and then give them a good polish. When pouring the gels I use a pipette and do not blow out the last bit of liquid. Finally, after pouring the separation gel I CAREFULLY add a smal ...

Jun 27, 2009    02:35 PM

Replied to topic in Cloning (Recombinant Gene Expression) forum   ligation troubleshooting

I would also recommend using Invitrogen's TOPO vectors but I'd suggest doing your PCR using a proof-reading polymerase and then ligating into the TOPO-blunt plasmid. But, you said that you already have the PCR product in a pGEM-T plasmid - When you remove it using EcoR I and Sal I it gives you a ...

Jun 27, 2009    02:04 PM

Replied to topic in Cell Culture and Tissue Culture forum   Problems with J774A.1 cells

Just to add that some people in our department are now using temperature sensitive TC plates which enable them to passage macrophages more gently. I don't remember the supplier's details (any ideas HeeHaw?) - just that the plates are quite new and their supply is not high enough to satisfy current d ...

Jun 24, 2009    04:48 PM

Replied to topic in Cell Culture and Tissue Culture forum   Flowcytometry of testicular cells

Are the cells you are FACS analysing fixed?

Jun 24, 2009    04:40 PM

Replied to topic in DNA (PCR, Real Time PCR) forum   Ladder-effect seen in my pcr images!

I hope it works out with the new primers. The only other thing I can think of is that certain wells in your PCR block are not getting up to 95 degrees as they should. Could there be dirt/dust etc in some of the wells? I think I'd run some samples on a different PCR machine to rule this out. Good ...

Jun 24, 2009    04:27 PM

Replied to topic in Cell Culture and Tissue Culture forum   Problems with J774A.1 cells

I know this might sound stupid, but are you sure you didn't use un-treated plates by mistake? Are the plates you refer to 10cm petri dishes? (as used for pouring LB plates etc). We have had this problem in the past. The student that stocked the tissue culture lab grabbed a number of packs of petri d ...

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