> We do some LOH as well as microsatellite instability. I have some
> pictures on my website at www.hci.utah.edu/groups/genomics/services.htm
> which show the expected results.
>
> I have looked at microsatellite instability using the markers Roche has in
> its kit, and although BAT25 and BAT26 are very sensitive for MIN, I don't
> think they would be appropriate for LOH because of the 1bp up and down
> ladder. The other primers should work fine.
>
> A lot of the trouble we have with LOH or MIN is template dependent:
>
> * Template prepared from archived tumor samples which have
> who-knows-what kind of junk in them. Try primers with short product
> <200bp, titrate the starting sample concentration to see how low you can
> go (to dilute any PCR inhibitors), and use a hot start enzyme--we like
> Life Tech's Taq Platinum.
> * Tumor samples with a high proportion of normal cells. Get someone
> really competent to microdissect the tumor slice so you get just tumor.
> For some tumors (prostate for example) you're just going to have to live
> with high normal concentration, so you have to do the experiment several
> times and do the math to see if you are seeing a statistically significant
> difference or not.
>
> Hope this is helpful,
>
> Linda
> ----------
> From: sharon.palmer@spectrum-health.org
> Sent: Wednesday, February 9, 2000 9:08 AM
> To: Recipients of ABRF List
> Subject: LOH
> Importance: High
>
> Hi all,
>
> I thought that someone in the group might be of some assistance. We are
> attempting to do a study concentrating on the loss of heterozygosity using
> microsatellite detection. We have tried the Roche kit and other specific
> labeled primers. The results are ambiguous at best.
>
> If any has a protocol, suggestion or has used the ABI rer/loh kit, please
> send a quick note.
>
> Thanks!!!!
>
> Sharon Keely Palmer
> Spectrum Health
> 616.356.4062
>
>
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