Cholesterol/Cholesteryl Ester Assay Kit - Detection (ab102515)

I'm glad you found the information of help.

I'd be very interested to know how you get on with the kit if you would mind letting me know. If you need any help with the kit please do not hesitate to contact us again.

Until then, I wish you all the best with your research.

Thank you for contacting us yesterday.

I now have some further information to share with you in regards to the Cholesterol/Cholesteryl Ester Quantitation Kit and Cholesterol/Cholesteryl Ester Detection Kit (ab65359 and ab102515 respectively).

As discussed over the phone, I have sought further information in regards to these kits and their suitability with egg samples. As mentioned on our product details page, this kit has as yet onlybeen tested for compatibility with mammalian samples. We have not tested egg yolk with either of thesekits. However, the source of this kit feels that there is no reason why it would not work. It would probably require some optimisation on your part to get the best results.

I looked at the publications we have on file for this kit and could not find any pertaining to egg yolk samples. It does not mean that it has not been done yet, just not published prominently. I would still suggest it should be suitable. However, we would not be able to guarantee it.

I hope this information has been of help. If you have any further questions, please do not hesitate to ask.

Thank you for your inquiry.

I can confirm that without the help of vacuum the drying process will be much longer.

But it is possible to do so by prolonging the air dry period at 50C.

Since this has not be experimentally tested by us, the length of the air dry incubation has to be determined by your customer.

I hope this information is helpful and wish your customer good luck with their experiments and their presentation.

Thank you for contacting us.

Yes, the kit is suitable for assay of mouse bile.

Please do not hesitate to contact us if you need any more advice or information.

Thank you for contacting us.

Indeed, for thequantification of cholesterol and cholesterol esters with have the kits ab65359and ab102515 available.ab65359uses both colorimetric and fluorometric detection, whereas ab102515 uses only colorimetric detection, but this assay can tolerate interferences from various samples significantly.

For best results we recommendextraction of lipids following theprotocol:

For cells or tissue samples, 106 cells or 10 mg tissue can be extracted with 200µl of chloroform: Isopropanol:NP40 (7:11:0.1) in a microhomogenizer. Centrifuge the extract for 5-10 minutes at 15,000xg. Transfer all of the liquid (organic phase), avoiding the pellet, to a new tube and air dry at 50ºC to remove chloroform. Put the samples under vacuum for 30 minutes to remove trace organic solvent. Dissolve dried lipids with 200µl of Cholesterol Assay Buffer by sonicating or vortexing until homogeneous (it is all right if the solution becomes cloudy). The extraction procedure can be scaled up if larger amounts of sample are required. Use 1-50µl of extracted sample per assay adjusting the volume to 50µl/well with Cholesterol Assay Buffer. For unknown samples it is recommended that different amounts are tested to ensure that readings are within the linear portion of the standard curve.

Unfortunately, we have not tested either kit without purification, so cannot make any further recommendations.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

1) A good stopping point would be after the samples are dried "(store at -20 °C until ready to proceed): Transfer the liquid phase to a new tube and air dry at 50°C then place samples under vacuum for 30 min to remove any remaining solvent." 2) Usually one does some 30-50 passes with a Dounce homogenizer- or you could try gently pipetting up and down (NO VORTEXING as this creates bubbles); or some sonication on ice if you do not have a homogenizer. Check a few microliters under the microscope to confirm you have good lysis of cells (if that is what you are working with). I hope this is helpful. Please contact us again if you have any further questions.