Antibodies, A Laboratory Manual, Second Edition, Edited by Edward A. Greenfield



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Labeling Antibodies with Cy5-Phycoerythrin

(Protocol summary only for purposes of this preview site)

Conjugates of the FRET dye Cy5-phycoerythrin (Cy5PE) with antibodies are relatively straightforward to make. The protocol does require synthesis of the Cy5PE tandem dye. Phycoerythrin can be purchased from multiple vendors. This type of conjugate is useful for immunofluorescence studies involving protein targets with low expression levels. Although the entire conjugation can be performed in a single day, there is an overnight stopping point (Step 8). When initially making Cy5PE derivatives, several different conjugates with varying ratios of Cy5 to PE should be made. These should be tested by conjugating to a well-characterized antibody. Absorbance spectra readings are a very worthwhile step to determine the quality of the Cy5PE label.

Antibodies: A Laboratory Manual, Second edition
Antibodies: A Laboratory Manual, Second edition
Antibodies: A Laboratory Manual, Second edition

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