Hi, I am culturing mammalian cells and would like to measure the glucose uptake. I have tried the LC-MS method. I am also aware of the GC-MS method using derivatization.
But I am wondering if there are any faster ways to do the measurement with less sample preparation?
Mrs/Miss Huang,
Please, consider my answer to the following discussion [A]; as well as the references mentioned, therein. The method shown, therein, is absolute from the perpective of chemometrics, which means that the correlation between experiment and theory according to your model formulas of quantification of the mass spectrometric outcome ''intensity'' shows coefficients up to /r/ = 1. The method is applicable to complex mixture of analytes at very low concentrations. It is already tested at pg.(mL)-1 and ng(L)-1 concentrations. Owing to the fact that the method od mass spectrometry operates at ultra-high resolving power; and, in parallel the discussed quantitative approach treats the intensity of the MS peaks per span of the scan time, there can be determined analytes via this approach almost without sample pretreatment.
Of course, there should be detailed more precisely from the perspective of the ''Analytical chemistry'' what is understood under the notion ''complex mixture of analytes.''
The method shown in [A] quantifies directly the analytes concentration in solution, as well.
[A] [https://www.researchgate.net/post/How_can_we_detect_VOCs_in_exhaled_breath_sample_using_ISQ_7000_single_quadrupole_GC-MS_system]
Hello, there were several possibilities for detection of glucose content.
Microbial biosensors, chemical titration, spectrophotometry, HPLC-RID etc.
All depends from analytical rage for more reliable answer provide information about glucose concentration that you want to detect.
It would be useful to choose a detector and separation technique after the estimation of the approximate concentration of glucose. As the authors indicated there are diverse measurement approaches for glucose. You need to narrow down the possibilities by the decision of the detection range of glucose in the cell culture medium. MS is compatible to quantify glucose levels as well, just after proper dilution and HILIC separation using proton acceptor resin such as amine and amide.
Emir...
Hi Luojiao,
do you rely on LC/ or GC/MS systems? Otherwise you can consider a variety of assay kits, using FACS or plate readers. For example, fluorescently labeled deoxyglucose, which will be taken up but not further metabolized, can be easily used.
Best,
Simon
Hi, Simon Lagies , I would like to try one of the simple assays for cell culture medium, such as something like blood glucose meters. My purpose is to rapidly calculate the uptake rate of glucose with a certain time period.
I recently found this product which seems a good solution to me. Would you recommend this?
https://www.kdbio.com/products/gluccell-meter-for-accurate-glucose-measurement-in-cell-culture-media/
Hi, Karen A. Darbinyan , Thank you for your answer. There is an estimate of glucose level around 20mM in the culture medium. But I have observed a severe signal suppression by using a HILIC-MS method. A parallel assay kit would be a good option to confirm the MS results.
Hi Luojiao. 20mM it is ppm level you can use various types of techniques.
1. Radioactivity
Intracellular accumulation of 3H-2DG6P
Requires multiple wash steps
Requires handling and disposal of radioactive materials
2. Fluorescence
Enzymatic detection of 2DG6P
Non-radioactive
Requires multiple sample preparation steps. Narrow window of detection.
3. Absorbance
Enzymatic detection of 2DG6P
Non-radioactive
Can detect very low 2DG6P
4. 2-NBDG
Intracellular accumulation of a fluorescently labelled glucose analog
Works well for imaging
Doesn't work well in plate formats
May not correctly reflect transporter activity.
5. Luminescence
Enzymatic detection of 2DG6P
Sensitive
Non-radioactive
Can detect very low 2DG6P with large assay window
Direct in-well detection with no wash steps
Scalable for high throughput
Not applicable for cell imaging
6. a method for measuring glucose uptake using radiolabeled 3-O-methylglucose;
7. a method for measuring glucose uptake using radiolabeled 2-deoxyglucose (2DG)
8. an improved method for measuring 2DG-uptake using an enzymatic, fluorometric assay.
9. LC-MS method of quantitative glucose and glycerol detection via enzymatic derivatization to glucose-6-phosphate and snglycerol-3-phosphate, respectively.
10. LC-MS method for assessment of glucose kinetics in vivo with D-[13C6]glucose as a tracer.
11. LC-MS method for Quantify 2-Fluoro-2-Deoxyglucose and Its Phosphorylated Metabolite for Determining Glucose Uptake.
12. LC MS Quantitative Analysis of Glucose by Cs+ Attachment.
Various methods with HPLC PDA, RID, FLU as well Glucose Uptake Colorimetric Assay Kit available.
hope this info will help you.
Good luck.
Please look at the following below links which may help you in your analysis:
https://www.genengnews.com/magazine/50/glucose-measurement-for-cell-culture/
https://www.scientific.net/MSF.941.2501
Thanks
I have no personal experience with the mentioned product! At least it is developed for cell culture and your concentration is within the given range. If you decide to buy it, you could test the system first by using fresh and cell-conditioned medium spiked with different amounts of glucose (optimally by not diluting the medium with water, so you could prepare some eppies, in which you evaporate your glucose solution and resuspend the pellet with medium).
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