hot start beads

[brett]

>Netters,
>We have used Invitrogen's HotStart Beads with wonderful results
>but they are VERY EXPENSIVE.  Cost is a major concern to us
>because we run LOTS of PCR reactions.  At the lowest cost we
>can purchase Invitrogen's beads (20,000+) it would still cost
>over $41 per 96 PCR reactions for the beads ALONE!!
>
>We are testing the use of paraffin wax (Aldrich) and mineral
>oil (used as a softener of the wax following the mixing of the
>limiting reagent and the reaction mixture) but have yet to hit
>upon a combination that we are happy with.  This approach
>requires a fair number of pipetting steps but the cost would be
>low.
>
>Invitrogen's beads have the Mg2+ dispersed throughout the bead.
> Is anyone in Netland willing to share their success in
>reproducing these kinds of beads?  I have thought maybe they
>could be produced by vigorously mixing molten wax and a
>solution of MgCl2 with a Polytron homogenizer.  Pipetting this
>mix onto a surface (eg. tin foil) might form a bead having the
>Mg2+ encapsulated in the wax.  Any thoughts on this?
>
>Whatever hot start technique you use, I would very much
>appreciate hearing about it.  If there is sufficient interest,
>I will summarize the replies and repost them to this bulletin
>board.
>
>Thank you in advance.
>
>Douglas Prasher
>U.S. Dept. of Agriculture
>Cape Cod , Massachusetts
>
>prasherd at delphi.com

After reading on this newsgroup (~2yrs ago?) about people making their own
beads, I thought I'd try it myself. Baxter Ameraffin LP tissue embedding
medium (cat#m7347) was melted (65oC), pipetted (15-20ul) onto a sheet of
aluminum, set, and stored at 4oC. Works great. In fact, a reproducible effect
in our lab is that our PCR's work better (ie greater yield) with these beads
than with PE's beads. I have no explanation, except there might be trace DMSO
in the paraffin from manufacture.


Brett Lindenbach
    
Program in Immunology                              
Washington University - St Louis                  
brett at borcim.wustl.edu                             

"I own my own pet virus. I get to pet and name her." - Cobain