User manual

Section 3- Applications
Note: This is unlike the other colorimetric assays on the NanoDrop 2000/2000c where it is recommended that
water be used as the blank.
• A standard curve can be generated using a minimum of two points which may include two standards or a
reference (Protein Pierce reagent/assay buffer only – no protein) and one standard.
• The multi-point curve capability allows for multiple replicates for up to seven standards. There is no set
order in which standards must be run.
• Additional standards may only be added before unknown samples are measured. Once the first sample is
measured, no additional standards or replicates may be added.
• A standard may be deleted at any time before a sample has been measured. Right-click the results table to
the far right to delete a standard or clear the entire standards table. Right-click a standard within the
replicates table under the graph display to delete individual standard replicates.
• Deleted standards cannot be added back or re-measured once the first sample is measured.
• In order to establish a new standard curve, a new workbook must be created. Colorimetric applications will
offer several options regarding the use of previous standard curves when appending sample data to
previous workbooks. It is recommended that the user follow the protein assay manufacturer’s guidelines for
generating standard curves before appending new data to a workbook.
Note: If selecting a previously saved workbook, all concentration calculations for newly measured
samples will be based upon the standard absorbance values saved in the workbook. Each
workbook will archive only one standard curve.
Making Protein Pierce 660 nm Assay Measurements
Sample and Standards Preparation
• Refer to the manufacturers’ guidelines and recommendations for sample preparation.
• The Zero Reference standard is a solution of the same buffer/dye reagent composition as the other
standards and samples but without any protein added.
• Prepare both standards and unknowns in the same manner. Be sure to use a diluent of the same pH and
ionic strength for all blanks, standards and unknown samples.
• Standards diluted from a stock must cover the expected range of the unknown samples. Pre-diluted
standard sets may be available from some manufacturers.
Procedure
1. Select the Protein Pierce 660 nm application from the main menu. If the wavelength verification window
appears, ensure the arm is down and click OK.
2. Enter the values for each standard concentration in the right pane table. The software allows for the
reference and up to 7 additional standards. The Reference and/or standards can be measured in replicates.
Note: The minimum requirement for standard curve generation is the measurement of two standards or
the measurement of the zero reference and at least one standard. It is recommended that
additional standards be included as necessary to cover the expected assay concentration range.
3. Select Add to report to automatically include all measurements in the current report. The default setting is
for all samples to be added to reports. The Add to report checkbox must be selected prior to a
measurement to save the sample data to a workbook.
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