© Thermo Hybaid. Issue 1.0, January 2002
64
In practice this difference in achieved temperature between different control systems is most
pronounced at the denaturation step. As a guide, we recommend that when transferring protocols
from a conventional thermal cycler to the Px2
In Situ
, the denaturation temperature is reduced by
3°C.
Programming
Programming and operation of the
In Situ
module are identical to the standard Px2 tube modules (see
Chapter 5) with the exceptions previously noted and also no heated lid operation is supported. For
simple fixed temperature incubations of the slides, manual control can be used (see section 4.7). For
more complex multi-step incubations or for temperature cycling reactions a protocol can be designed
and run using Simulated Slide Control (see Chapter 6).
Humidity Chamber
The polycarbonate Humidity Chamber has been designed to reduce drying out of the sample on the
slides. For accurate temperature performance, the
In Situ
module should be used with the Humidity
Chamber in place and the lid closed.
•
Fill the reservoirs with water.
•
Pre-heat the block with the Humidity Chamber in place. This allows the reservoir to warm up so
that evaporation of the solution can begin. Alternatively, preheat the reservoir solution to the
incubation temperature required.
•
For thermal cycling with extended/high temperature incubations, it is important to seal the reagents
under the cover slip. Thermo Hybaid recommends the use of EasiSeal sealing frames. These are
available in three sizes (25
µ
l - HB-OS-SSEZ1E, 65
µ
l - HB-OS-SSEZ2E, and 125
µ
l - HB-OS-
SSEZ3E). They will completely seal the specimen area to prevent drying.
NB:
If a petroleum based rubber compound is used for sealing cover slips in place, the compound
should be allowed to dry out completely before use with this system. If complete drying is not
allowed, the solvent vapour will damage the surface of the Humidity Chamber.
More information on the use of In Situ applications using Thermo Hybaid equipment can be
obtained from the Thermo Hybaid In Situ Hybridization Guide, which is available free upon
request.
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