( water into the wells in approximately ...)PathCheck Applied to Measurement of Protein Solutions in the SpectraMax Plus Microplate Spectrophotometer (MaxLine Application Note #26)

Step 2 Put the plate into the drawer of the SPECTRAmax PLUS, then click the Read button in SOFTmax PROs toolbar to make the pre-read measurements.

Step 3 Remove the plate, pour out the water and blot the plate dry.

Making pathlength-corrected absorbance measurements
Note: If you have elected to use the pre-programmed Water Constant, you may skip Step 1.
Step 1 Place a clean quartz cuvette containing your sample buffer or distilled water into the SPECTRAmax PLUS cuvette port.

Step 2 Transfer aliquots of your samples and blanks into their designated wells in the microplate. Accurate pipetting is not necessary, though the volumes should between 100 L and 300 L for best pathlength-corrected results.

Step 3 Put the plate into the SPECTRAmax PLUS drawer, then click the Read button in SOFTmax PROs toolbar to make the absorbance measurements. (The cuvette will be read in the near-infrared at the same time the plate is read, and its absorbance values used in the PathCheck calculations.)

To avoid evaporation error, make the readings within a few minutes of putting the samples in the microplate. If the readings must be delayed, cover the plate with an adhesive seal. Remove the seal immediately prior to reading the plate.

Storing and re-using pre-read data
Quartz plates are so uniform that it is not necessary to subtract the plate background on a well-by-well basis. Instead, you can pre-read one plate, then store the pre-read data in a SOFTmax PRO file for subsequent re-use, being careful to save the data under a different file name
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