I am trying to perform osteoblast quantification using H&E. I have been reading papers (like this one: Article Extracellular matrix with defective collagen cross-linking a...
)where measurements like osteoblast surface or osteoblast number per bone surface are derived from H&E images taken from the proximal tibia, distal femur etc. as the ASBMR outline for trabecular bone analysis.
How are these counts performed? Are the OBs thresholded using a purplish range of colours and then specified a certain maximum distance from the trabecular bone surface?
If this is the method, are they all assumed to be OBs and not OCs or other bone-lining cells?
I can quantify my OCs using TRAP staining quite nicely, however I would really like to also use H&E to quantify OBs as well.
I am unsure of the identification and quantification of OBs in H&E images.
Any help would be much appreciated, thanks very much.
Flynn
Dear Flynn,
You can definitely measure osteoblast parameters on H&E stained sections. I usually count the number of osteoblasts on bone perimeter and osteoblasts surface. This is done by identifying osteoblasts mostly through morphology, I see very difficult to find semi-automatic ways using threshold.
Osteoblasts are cuboidal cells found on bone trabeculae, they are never alone so you will never find only one osteoblast and so you cannot confuse them with osteoclasts.
If you use TRAP histochemistry with a very light hematoxylin counterstain, you can also easily identify osteoblasts, usually close to osteoclasts or on the other side of the bone trabecula.
I usually don't count bone lining cells, as they usually do not reflect active bone formation. But this is up to you, if you start counting them, then you need to do it for all your samples.
Ok Biagio I'll take this into consideration. Thanks so much for your help.
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