The sample volume range on that particular microplate reader was 200 uL to 300 uL. Any of those volumes within that range would have worked. If 300 uL was added, it would basically be a full well. We just chose 250 uL because it was in the middle of that range.
From what I can tell the model is a Spectramax M2e Microplate Reader, and the brand is Molecular Devices. Resellers like Marshall Scientific also sell these plate readers.
Small bubbles at the top of the liquid line shouldn't impact the absorbance, but if you have a super bubbly solution, it would likely affect the absorbance.
Hello, I am very new for this subject, could you please tell me which solution you took as a sample( blue sample ) or how to prepare the blue sample ? And how can I calibrate the absorbance microplate reader easily and which solution I need. I will be helpful if you give me the solution. Thanks
Hello! The solution for this video demo was blue food coloring in water. We made a stock solution of dark blue food colored water and then did simple dilutions for the solutions to make a standard curve. Please note your experiment will require a different solution for making the standard curve. Different molecules absorb at different wavelengths so it’s super important to make your standard curve with a solution specific for your experiment. For example if your experiment uses a buffer, you need to calibrate the microplate reader with the same buffer solution. Make dilutions with known concentrations of your compound of interest to make the standard curve.
@@musicgirl4229 Thank You so much. Could I use pH 4/7 or 10 as a sample solution ? Because all these solution I have are colored solution. So, can I use this buffer as sample solution for experiment?
You can do an absorbance spectrum to find the maximum absorbance - the maximum wavelength will have the most sensitivity and thus is the best to use to obtain the most optimal results. In general, the solution will absorb at the complementary color's wavelength. In the video, the color of the solution was blue, thus the solution absorbed light around the wavelength for the color orange.
