Glucose Assay Kit (ab65333)

The signal contributed by the phenol red in 1ul samples will be insignificant compared to the signal generated by the glucose in your samples. You should include a control consisting of 1ul of culture medium and subtract the absorbance or fluorometric value of that control from the values of your experimental samples.

Thank you for contacting us. Yes, peroxide is known to interfere with this Glucose Assay Kit. I hope this helps, please let me know if you need any additional information.

This kit is only specific for D-glucose.

I have been in contact with the lab and unfortunately, we have not tested whether the addition of acids such as H2SO4 or PFA will interfere with the assay.

They have suggested that in this situation, perhaps it is best if you deproteinise your samples to remove most proteins/enzymes which would constitute the “biohazard” in your samples.

Alternatively, would 10% SDS be suitable for inactivating your samples? Although we have not tested this, itght work since it has been used in other assays with similar chemistry.

We have not tested glucose in purified polysaccharide samples. The first step of the reaction is specific for glucose, so we expect this to work. It's possible that there will be some background, but this can be corrected for by subtracting the background control values (see page 8 of the protocol).

The main difference between these two glucose assays is that in ab65333 the probe reacts with hydrogen peroxide and gets oxidized to produce colored compound detectable at 570nm or at Ex/Em 535/587nm. In ab102517, the probe reacts with NADH (produced during glucose dehydrogenation in the first step) and forms a colored product measured at 450nm.

Assay ab65333 can measure 1uM-10mM glucose. The additional sensitivity in ab65333 is achieved in the fluorometric measurement of the assay. Assay ab102517 can measure concentrations within the range of 20uM-10mM.

I suggest exchanging the medium before collecting samples. That is, culture the cells in medium with a glucose/energy source and then exchange the medium with glucose-free medium such as Krebs-Ringer-Phosphate-HEPES (KRPH) buffer for example, and then run the glucose assay using the KRPH buffer. All the glucose in the buffer will be from the cells. There might be need for stimulation of glucose transporter for the cells to release glucose into the buffer. I would strongly suggest referring to publications on these types of experiments for protocols.

Thank you for your enquiry.


Answer 1: Yes, the minimal serum volume for this assay is 0.5 µl. The lower detection limit of this assay is 1 µM. That does not mean that you have to start with a particular conc of the sample. More specifically, you would not know the conc of glucose to begin with.



Answer 2:

In the protocol step mammalian serum consists of 0.1-6 mM triglyceride. This is the conc of triglyceride, not serum. One can use a minimum of 1 µl and a max of 50 µl for this assay.

This kit would be used to measure glucose in the cell culture supernatant. The number of cells in culture would have an effect on the glucose consumption, but the amount of time the cells are cultured and the specific cell line you are testing for will also have an effect on glucose levels.

Therefore, it is difficult to provide precise details of the number of cells you should use. I can suggest growing the cells as normal, taking samples at various time points. In the protocol, we suggest testing several doses of your sample to make sure the readings are within the standard curve range. You may need to dilute them in order to ensure they are within range, in which case you will need to take this into account when calculating the glucose concentration from the reading obtained.

I am sorry it is difficult to be more precise than this for individual experiments, but I hope this will be helpful.

Thank you for your enquiry which has been forwarded to the scientific support team.

I can confirm I would suggest Glucose Detection Kit ab102517or ab65333 Glucose Assay Kitswill be suitable for detection andquantification of glucose in the culture samples. I have put information below regarding the sensitivity and detection range of the kits, I can suggest to review this to decide which kit is best for your experiments (I am sorry we do not have concentration expressed as a percentage, I am not sure what this would be).

ab102517 has a senstivity of > 0.02 mM, range 0.02 mM - 10 mM
https://www.abcam.com/Glucose-Assay-Kit-reducing-agent-compatible-ab102517.html (or use the following: https://www.abcam.com/Glucose-Assay-Kit-reducing-agent-compatible-ab102517.html).

ab65333 has a sensitivity of 1 µM, range 1 µM - 10000 µM
https://www.abcam.com/Glucose-Assay-Kit-ab65333.html (or use the following: https://www.abcam.com/Glucose-Assay-Kit-ab65333.html).

I am sorry we do not sell directly to Russia. However, we do have an official distributor in Russia, Almabion. I can recommend to contact them directly to request a price quote:

ALMABION
ul. Vladimira Nevskogo d.59, k.166
Voronezh
394005
Russian Fed

Te l+7 4732 734828
Websitehttp://www.almabion.ru
Fax+7 4732 734828
mailto:Emailinfo@almabion.ru

I hope this will be helpful to you. Please do not hesitate to contact me if you have any further questions.