Standardized measurements and differential spectroscopy in microplates.
Microplates (MPs) are excellent devices for the parallel processing of multiple samples for the spectroscopic analysis of chromophores and turbidity, for luminometric measurements, for cell culture applications, or simply for sample storage, library organization, and other high-throughput (HTP) processes. Disadvantages include an ill-defined pathlength, meniscus formation, evaporation, and cross-contamination. Here, we have developed a novel MP and lid system which can serve to minimize these drawbacks. Cup-like lids are inserted into MP wells. Thereby, liquid is pushed aside. The flat bottoms of the cup-like lids guarantee a planar interface and a defined pathlength. In addition, the devised MP system allows for differential spectroscopic analysis of multiple samples comparable to measurements in tandem cuvettes. This was shown by the investigation of the binding of reduced nicotinamide adenine dinucleotide to dihydrolipoamide dehydrogenase. The MP lid system described offers a low-cost solution for standardized spectrophotometric quantitations in any solvent compatible with the MP/lid material. In addition to the system's suitability for routine MP application, it should be advantageous as a simple and noninvasive method, i.e., no labeling and immobilization of analytes is required for detection of the interaction of molecules, for various HTP applications and drug screening purposes.
Geiger
G
G
Laboratory of Biochemistry, Institute for Interfacial Engineering, University of Stuttgart, Fraunhofer IGB, Nobelstrasse 12, Stuttgart, D-70569, Germany.
Bernhagen
J
J
Wagner
E
E
Bisswanger
H
H
Brunner
H
H
Vitzthum
F
F
D001665:Binding Sites; D008058:Dihydrolipoamide Dehydrogenase / Q000032:analysis; D005470:Fluorometry; D008027:Light; D009243:NAD / Q000032:analysis; D013048:Specimen Handling / Q000295:instrumentation*; D013057:Spectrum Analysis / Q000295:instrumentation* / Q000592:standards*
D009243:NAD; D008058:Dihydrolipoamide Dehydrogenase