Atto 465 Derivative Is a Nuclear Stain with Unique Excitation and Emission Spectra Useful for Multiplex Immunofluorescence Histochemistry

Joshua T. Dodge; Andrew D. Doyle; Ana C. Costa-da-Silva; Christopher T. Hogden; Eva Mezey; Jacqueline W. Mays

doi:10.1369/00221554211064942

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Atto 465 Derivative Is a Nuclear Stain with Unique Excitation and Emission Spectra Useful for Multiplex Immunofluorescence Histochemistry

Journal article

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Atto 465 Derivative Is a Nuclear Stain with Unique Excitation and Emission Spectra Useful for Multiplex Immunofluorescence Histochemistry

Joshua T. Dodge, Andrew D. Doyle, Ana C. Costa-da-Silva, Christopher T. Hogden, Eva Mezey and Jacqueline W. Mays

The journal of histochemistry and cytochemistry, Vol.70(3), pp.211-223

01/07/2022

DOI: 10.1369/00221554211064942

PMCID: PMC8832627

PMID: 34994225

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Abstract

Multiplex immunofluorescence (mIF) is an effective technique for the maximal visualization of multiple target proteins in situ. This powerful tool is mainly limited by the spectral overlap of the currently available synthetic fluorescent dyes. The fluorescence excitation wavelengths ranging between 405 and 488 nm are rarely used in mIF imaging and serve as a logical additional slot for a fluorescent probe. In the present study, we demonstrate that the addition of 2,3,4,5,6-pentafluoroaniline to Atto 465 NHS ester, creating Atto 465-pentafluoroaniline (Atto 465-p), generates a bright nuclear stain in the violet-blue region of the visible spectrum. This allows the 405 nm excitation and emission, classically used for nuclear counterstains, to be used for the detection of another target protein. This increases the flexibility of the mIF panel and, with appropriate staining and microscopy, enables the quantitative analysis of at least six targets in one tissue section. (J Histochem Cytochem XX: XXX-XXX, XXXX)

Cell Biology

Life Sciences & Biomedicine

Science & Technology

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Title: Subtitle: Atto 465 Derivative Is a Nuclear Stain with Unique Excitation and Emission Spectra Useful for Multiplex Immunofluorescence Histochemistry
Creators: Joshua T. Dodge - National Institute of Dental and Craniofacial Research
Andrew D. Doyle - National Institute of Dental and Craniofacial Research
Ana C. Costa-da-Silva - National Institute of Dental and Craniofacial Research
Christopher T. Hogden - National Institute of Dental and Craniofacial Research
Eva Mezey - National Institute of Dental and Craniofacial Research
Jacqueline W. Mays - National Institute of Dental and Craniofacial Research
Resource Type: Journal article
Publication Details: The journal of histochemistry and cytochemistry, Vol.70(3), pp.211-223
DOI: 10.1369/00221554211064942
PMID: 34994225
PMCID: PMC8832627
NLM abbreviation: J Histochem Cytochem
ISSN: 0022-1554
eISSN: 1551-5044
Publisher: Sage
Number of pages: 13
Grant note: ZIC DE000750 / NIDCR Imaging Core
Language: English
Date published: 01/07/2022
Academic Unit: Endodontics
Record Identifier: 9984367728902771

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