Hi,
I'm looking for a negative control for osteocalcin expression (ELISA, PCR) - but seems to be very widely expressed...
https://www.ncbi.nlm.nih.gov/gene?Db=gene&Cmd=DetailsSearch&Term=632#gene-expression
Does anyone know or have tried a tissue or cell line that could be used as a negative control?
Thank you!
Sophie
what have you looked at so far- perhaps try some lymphocytes or other WBCs
Nothing yet but just RNA seems to be widely expressed so wondered if an solid negative controls were known, just to save any time - cheers I'll see if any WBCs are available in our lab :) going to try a few, will let you know!
If osteocalcin is made by osteoclasts and is found in bone and dentin, which are mesenchymal, I would choose a cell line that is endodermal (e.g., gut and lung) or ectodermal (e.g., skin and neural) as negative controls.
It is a worthy of note that almost anywhere in the body where a long-standing dystrophic calcification occurs may lead to heterotopic bone formation, suggesting that other cell types have the genetic information for turning on osteocalcin formation.
You can isolate lymphocytes from peripheral blood by Hi se(Hi Media), do a viable cell count by Trypan blue and dilute the suspension in cell culture medium for seeding as control cells
A quick comment on Ashraf's post- Osteoblasts, not Osteoclasts, make bone. Osteoclasts are glorified macrophages and are thus hematopoietic cells. Also, ectopic ossification likely arises from tissue resident mesenchymal cells (vascular pericytes, which are everywhere) and is not really a reflection of transdifferentiation of non mesenchymal cells but rather aberrant signalling through BMPs or similar factors. Also curious if you've seen osteocalcin protein in any of your non osteoblast cells or just RNA? I'd always thought that OC expression was pretty tightly restricted to osteoblastic cells.
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