I am going to start a feeding experiment on sheep under heat exposure.
I want to determine the circulatory plasma insulin and thyroxine hormone. It would be highly appreciated if anybody can help with a detailed laboratory procedure for these two hormones. The procedure can be ELISA or any other non-isotopic technique.
You need to decide whether you are going to estimate by ELISAor other techniques, accordingly commercial kits are available (species specific) where Ab tagged plates will be provided with the detailed stepwise procedure.
You can probably use an ELISA kit for both. I usually start with biocompare.com. It's basically a search engine for all sorts of products for research, and will let you narrow your search by species. The kit will come with a protocol.
http://www.biocompare.com/Assay-Kits/
Thanks Nicole, I will go through your link and will see what are the equipments we need for running that kit and the availability...
You should probably only need pipets and a spectrophotometer, maybe some chemicals and a deionized water system.
As Ahmen stated, T3 & T4 are not species specific so any commercially available kit would do. The only issue though is the sensitivity of the kit. Sheep have a far lower concentration of both endocrines in their system compared to humans, so make sure your kit is reasonably sensitive. I found the human competitive ELISA kits from Abnova Corporation very effective with a sensitivity of 0.05pg/mL, but there are many manufactures of kits in this world. Just check your sensitivity!
An side question though Rahman, why are you focusing on T4? Triiodothyronine (T3) is the active form and variations in its concentration would give you a better indication of metabolic response to heat stress. Another interesting investigation may be changes in Reverse T3 due to heat stress. Just considerations!
For insulin you can use Mercodia Ovine Insulin kit (ELISA). The procedure is very simple. You just need a plate shaker and a plate reader. I've no idea about the thyroxin. The big issue is the cost of each package, because with each package you can measure just 84 samples alone and 42 samples in duplicate!
Hope this helps,
Ehsan
As a reply to Mark Barnett, till now our lab solely worked on digestive and metabolic response in sheep and nothing about metabolic hormones. So, I am trying to introduce and relate our research with metabolic hormones. It will be highly appreciated if you could suggest any paper related to metabolic hormones in relation to heat stress of ruminants that can potentially enrich my understanding.. Thanks a lot..
You can look for an immulite machine to run insulin samples . This method is cheaper and faster than elisa. However, you need to find someone who has that machine
hi ..i am experianced in CLIA method ,its really good.both insulin and free t3,freet4 and Tsh like immune assays are good in that.
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