( cubator (for test tube/cuvette method) ...)Microdetermination of Phosphorus Using the SpectraMax Plus Microplate Spectrophotometer: Choice of Microplate, Cuvette, or Test Tube Assay Formats (MaxLine Application Note #24)

8. Sulfuric acid, reagent grade

9. Ammonium molybdate, ACS reagent grade

10. Ascorbic acid, ACS reagent grade

CHEN et al METHOD
Setting up the instrument and the software
Step 1 Using SOFTmax PRO, select a Cuvette Set (test tube/cuvette method) or a Plate Section (microplate method). If necessary, create a new Cuvette Set or Plate Section by selecting New Cuvette or New Plate from the Experiment menu.

Step 2 Set up the Instrument Settings dialog box as shown in Figure 4 for a cuvette (upper) or for a plate (lower). Select to perform an end-point read at 820 nm. DO NOT SELECT PATHLENGTH CORRECTION (i.e., leave the PathCheck checkbox unchecked). The phosphomolybdate absorption band extends into the near IR and interferes with the measurements needed for pathlength calculation. It is not necessary to preread the microplate because polystyrene contributes negligible absorbance at 820 nm.

Step 3 Create a template showing where standards and unknowns will be located in the Cuvette Set or the Plate Section.

Step 4 Click the Reduction button in the Cuvette Set or the Plate Section toolbar to display the Reduction dialog box. Set the wavelength combination to L1 and the data mode to Absorbance as shown in Figure 5.

Step 5 If using the microplate method, you have the option of performing the incubation step in the microplate reader or in an external incubator. If the plate is to be incubated in the plate reader, set the incubator to 37 C (using either the front panel of the instrument, or SOFTmax PROs incubator (thermometer) button). If using tubes (or a microplate, if you dont want to use the instrument to incubate it), check that a 37 C incubator is available for them.

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