NIST Spectroscopic Measurement Standards
Abstract
Ultraviolet (UV) absorbance measurements provide a rapid and reliable method to determine protein concentrations. The National Institute of Standards and Technology (NIST) has developed Standard Reference Material (SRM) 2082as a pathlength standard for UV absorbance measurements for use with the new generation of microvolume spectrophotometers and short-pathlength cuvettes. Short pathlengths are used with high-concentration targets to ensure that absorbance values are within the optimal range. The short-pathlength instruments and cuvettes also reduce the required volumes to conserve valuable samples. The authors compared the results obtained with high-quality dual-beam spectrophotometers and short-pathlength cuvettes to the results obtained from a microvolume spectrophotometer and a microvolume plate reader. SRM 2082 can be used to accurately calculate pathlength values, thereby increasing the accuracy in subsequent measurements using the short-pathlength cuvettes and microvolume absorbance instruments. RM 8671 (reference material, the NISTmAb) can then be used to ensure the accuracy and reproducibility of protein concentration measurements by providing an industrially relevant reference material, a well-characterized monoclonal antibody.
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Peer-Reviewed
Submitted: Jan. 22, 2018
Accepted: Feb. 16, 2018
About the Authors
Kenneth D. Cole*is group leader bioassay methods, [email protected]; Paul DeRose, Brian Lang, John Schiel, and Aaron Urbas are research chemists; Hua-Jun He and Erica V. Stein are research biologists; Evelyn Solis is summer student; and Steve Choquette is director, Office of Reference Materials—all are at NIST.
*To whom all correspondence should be addressed.