Can I quantify D-Lactate levels?
No, the kit is used for the quantification of L-Lactate levels.
How stable is the Working solution?
You can’t store the Working solution. Please prepare the Working solution prior to use. Please protect from light because the Working solution is light-sensitive. The Working solution is stable for 4 hours at room temperature with protection from light.Once the Working solution is exposed to light, the color of the solution becomes from red to orange. It caused to increase background.
Can I measure samples that contain a reducing agent?
You can’t measure an accurate Lactate level because WST dye interferes with the reducing agent. If you add test material which possesses reducing properties, please prepare a few wells that only contain the medium without cells and only test material for background control. Please subtract the background from the absorbance of standard curve or samples
Are there protocols for measuring intracellular lactate level?
(1) Collect cells (1 × 105 cells * 1) in a microtube.(2) Centrifuge at 300 × g for 2 minutes and remove supernatant.(3) Add 300 μl of cold PBS and suspend by pipetting. Centrifuge at 300 × g for 2 minutes and remove supernatant.(4) Add 300 μl of 0.1% Triton-X * 2 and vortex for 1 minutes to prepare for cell lysate.(5) Centrifuge at 8000 × g for 5 minutes and collect supernatant.(6) Deproteinize the solution prepared at step (5) with an ultrafiltration membrane filter (fraction molecular weight: 10 K) * 3 to obtain a measurement sample.* 1 For HeLa cells, more than 1 × 105 cells are necessary to detect lactate level over 0.02 mmol / l.* 2 When SDS is included in the cell Lysate, color development can be inhibited. Please avoid using buffer containing SDS.* 3 Since endogenous Lactate dehydrogenase (LDH) causes background please remove LDH by deproteinization treatment.
How many samples can I measure?
* The number of samples that can be recorded when the standard curve and sample is measured in triplicates.
*A plate layout for Lactate standard solution and sample
I do not have a 450 nm filter. What other filters can I use?
You can use filters with an absorbance between 450 nm and 490 nm. However, the absorbance value can become lower if the sample is not measured at 450 nm.
Our samples did not change in color, are there any reasons for this?
Lactate level contained in the sample may be lower than the detection limit (0.02mmol/l) that can be measured using this kit. If the lactate level is below 0.02mmol/l, please consider other measurement method such as LC-MS.If you dilute the sample, lactate level contained in the diluted sample may be lower than 0.02mmol/l.Please lower the dilution ratio and adjust the concentration of the sample over the detection limit that was measured using this kit.