Sensitivity Detection limits using proteins – EkkO – Spectroscopy – Application Note 32

Abstract

In this note, we present CD measurements of ()-lactoglobulin, Cytochrome C, Hemoglobin and Lysozyme with variable well volumes and concentrations to address the sensitivity of the EKKO; CD Microplate Reader for CD measurements. We show the EKKO; CD Microplate Reader is linear for the determinations of absorbance and CD up to a value of ~ 1.2 equating to a protein concentration of 50 to 100 g/ml for a helical protein.

Introduction

Circular dichroism (CD) refers to the differential absorption between left and right circularly polarized light is commonly used for chiral analysis. Generally, it is used for determining enantiomeric purities in asymmetric syntheses and assigning the secondary structures of proteins, both of which require the ability to do the measurements in a high-throughput
fashion.

Recently these determinations have been expedited by the addition of robotic liquid handling systems coupled to a conventional CD spectrometer. However, transferring individual samples and cleaning the cuvette between measurements makes CD measurements with traditional CD Spectropolarimeters a time-consuming process…

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