I have recently purchased the
ImmPRESS® Duet Double Staining HRP/AP Polymer Kit (anti-mouse IgG-brown, anti-rabbit IgG-red) Cat no: MP-7724 .
My aim is to detect macrophages in breast carcinoma, where the brown (CD163, mouse monoclonal) stains the cytoplasm while the red stains the nucleus (cMAF, rabbit polyclonal) of the macrophages. I have tested both my primary antibodies with DAKO DAB kit, and the antibodies work perfectly.
However, i can't seem to get any result when i used the double stain kit. When i dehydrated and cleared the slides, the red stain seems to wash of. If i do not dehydrate, the red stain is too strong and gave a lot of non-specific staining. Worse part is , it does not even stain the target intended, which is the nucleus. I have tested this kit with Ki67 (mouse) and HER2 (rabbit) antibodies, it seems to work fine. Where did i go wrong?
Can contact the vendor
Follow the instruction from the kit carefully
Focus on washing steps and dilution of abs
You may want to restructure your question
"detect macrophages in breast carcinoma" may provide more info
also can post a pic of the stained cells
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