
CLONEGENE
A forum for further information on CLONEGENE products and user feedback.
Sunday, August 2, 2009
The detection of HuR in tumor tissues.

Friday, July 31, 2009
Western blotting with an anti-HuR monoclonal

For further information contact clongene or email info@clonegene.com.
Friday, July 24, 2009
Monoclonal antibodies to Dicer - CG006 and CG016

CG006
CG006 is available as a purified monoclonal is very heat stable and routinely shipped at room temperature in a proprietary stabilizer solution. But it is convenient to keep it in fridge for short-term use (weeks) or in freezer for long-term use (months). For immunochemistry or immunofluoresence we use 1/500 to 1/2000 dilutions and use standard anti mouse ABC peroxidase detection systems from Vector labs. For western blots we use similar dilutions and the standard anti – mouse systems using ECL detection.
CG016
Cg016 is currently available as a hybridoma and is also heat stable and shipped at room temperature. It is convenient to keep it in fridge for short-term use (weeks) or in freezer for long-term use (months). For detection of Dicer on paraffin sections, we use dilution of 1/50 to 1/200 and the vector labs ABC elite detection kit. The sections are treated with RETRIEVIA as discussed in earlier post.
Labels:
ABC,
Antibody,
Dicer,
MicroRNA,
Pleuropulmonary Blastoma
Wednesday, July 22, 2009
RETREVIA - A new antigen retrieval reagent for IHC

However, this fixation method cross-links proteins and thus these sections do not work well in standard immunohistochemical assays. In most cases the epitope recognized by a particular the cross-linked residues obscure antibody and little reactivity is possible. To reactivate the protein antigen, the sections are heated in a buffer, which removes the crosslinks. Some protocols recommend steam heat some recommend microwave. Clonegene has now launched a new retrieval reagent called RETREVIA. The above picture shows the strong reactivity of a lung cancer section with our anti-HuR antibody (CG001) after treatment with RETREVIA .
Below we summarize the key steps that we recommend to give superior staining with our antibodies
1. Deparaffinize sections in 2 changes of xylene, 5 minutes each.
2. Hydrate in 2 changes of 100% ethanol for 3 minutes each, 95% and 80% ethanol for 1 minute each. Then rinse in distilled water.
3. Pre-heat steamer or water bath with staining dish containing RETREVIA (dilute 10 ml of RETREVIA to 1 Litre of distilled water) until temperature reaches 95-100 °C.
4. Immerse slides in the staining dish. Place the lid loosely on the staining dish and incubate for 30 minutes (however this incubation time may depend on the nature of the sections and should be determined by user).
5. Turn off steamer or water bath and remove the staining dishes to room temperature and allow the slides to cool for 20 minutes.
6. Rinse sections in PBS for 2x2 min.
Proceed to standard immunohistochemistry protocol, we recommend Vectors labs ABC elite staining kit.
RETREVIA also works well in automated staining platforms.
For sales or other information contact Clonegene
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