Absorbance detection measures the ability of a sample to absorb light of a specific wavelength. It was the first detection mode available in microplate readers more than 3 decades ago. Unlike other detection modes, absorbance is a measure through the sample. Absorbance microplate readers determine the amount of light absorbed by a sample by quantifying how much of the light the sample was exposed to is transmitted through it. Hence absorbance readers require a light source illuminating the sample, an optical filter or monochromator, selecting a specific wavelength, and a light detector positioned on the opposite side measuring the amount of transmitted light. In spectrometer-based readers a selection of the wavelength is not necessary as the whole spectrum (usually from around 220 to 1000 nm) is acquired.
Absorption of light is often referred to as the Optical Density (OD) and is a function of the concentration of a sample. Consequently, absorbance detection can be used for quantification purposes.
Common applications include ELISA assays, bacterial growth, enzyme activity assays, and DNA/RNA and protein quantification. Moreover, conventional colorimetric analyses have been adapted to the microplate format and can be quantified by plate readers.
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