TY - JOUR
T1 - SeqStain is an efficient method for multiplexed, spatialomic profiling of human and murine tissues
AU - Rajagopalan, Anugraha
AU - Venkatesh, Ishwarya
AU - Aslam, Rabail
AU - Kirchenbuechler, David
AU - Khanna, Shreyaa
AU - Cimbaluk, David
AU - Kordower, Jeffrey H.
AU - Gupta, Vineet
N1 - Funding Information:
We thank Liudmila Romanova and Diptaman Chatterjee in Kordower laboratory at Rush University Medical Center and the Center for Advanced Microscopy at Northwestern University for help with tissue staining and image analysis, and all past and current members of the Gupta laboratory for their technical help and suggestions. This work was supported in part by funding from the Oppenheimer Family Foundation, Bears Care, the Department of Internal Medicine at Rush University Medical Center, and by the National Institutes of Health (grants R01DK107984, R01DK084195, and R01CA244938 to V.G.). Conceptualization, V.G. A.R. and I.V.; methodology, A.R. I.V. R.A. D.K. J.H.K. and V.G.; investigation, A.R. I.V. R.A. and S.K.; software, A.R. S.K. and D.K.; supervision, V.G.; writing – original draft, A.R. I.V. R.A. D.K. S.K. D.C. and V.G.; writing – review & editing, A.R. I.V. J.K. and V.G.; funding acquisition, V.G. A.R. and V.G. are inventors on a patent application filed by Rush University Medical Center related to these studies. V.G. is a co-founder of 149 Bio, LLC and Spatialomyx, LLC that is licensing this intellectual property and has significant financial interest in it. V.G. is also an Officer and a Board member of 149 Bio, LLC and Spatialomyx, LLC. The remaining authors declare no competing interests.
Funding Information:
We thank Liudmila Romanova and Diptaman Chatterjee in Kordower laboratory at Rush University Medical Center and the Center for Advanced Microscopy at Northwestern University for help with tissue staining and image analysis, and all past and current members of the Gupta laboratory for their technical help and suggestions. This work was supported in part by funding from the Oppenheimer Family Foundation , Bears Care , the Department of Internal Medicine at Rush University Medical Center , and by the National Institutes of Health (grants R01DK107984 , R01DK084195 , and R01CA244938 to V.G.).
Publisher Copyright:
© 2021 The Authors
PY - 2021/6/21
Y1 - 2021/6/21
N2 - Spatial organization of molecules and cells in complex tissue microenvironments provides essential organizational cues in health and disease. A significant need exists for improved visualization of these spatial relationships. Here, we describe a multiplex immunofluorescence imaging method, termed SeqStain, that uses fluorescent-DNA-labeled antibodies for immunofluorescent staining and nuclease treatment for de-staining that allows selective enzymatic removal of the fluorescent signal. SeqStain can be used with primary antibodies, secondary antibodies, and antibody fragments to efficiently analyze complex cells and tissues. Additionally, incorporation of specific endonuclease restriction sites in antibody labels allows for selective removal of fluorescent signals while retaining other signals that can serve as marks for subsequent analyses. The application of SeqStain on human kidney tissue provided a spatialomic profile of the organization of >25 markers in the kidney, highlighting it as a versatile, easy-to-use, and gentle new technique for spatialomic analyses of complex microenvironments.
AB - Spatial organization of molecules and cells in complex tissue microenvironments provides essential organizational cues in health and disease. A significant need exists for improved visualization of these spatial relationships. Here, we describe a multiplex immunofluorescence imaging method, termed SeqStain, that uses fluorescent-DNA-labeled antibodies for immunofluorescent staining and nuclease treatment for de-staining that allows selective enzymatic removal of the fluorescent signal. SeqStain can be used with primary antibodies, secondary antibodies, and antibody fragments to efficiently analyze complex cells and tissues. Additionally, incorporation of specific endonuclease restriction sites in antibody labels allows for selective removal of fluorescent signals while retaining other signals that can serve as marks for subsequent analyses. The application of SeqStain on human kidney tissue provided a spatialomic profile of the organization of >25 markers in the kidney, highlighting it as a versatile, easy-to-use, and gentle new technique for spatialomic analyses of complex microenvironments.
KW - Spatialomics
KW - antibody DNA conjugates
KW - cellular neighborhoods
KW - multiplex immunofluorescence
KW - relationship maps
KW - sequential staining
KW - spatial pathology
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U2 - 10.1016/j.crmeth.2021.100006
DO - 10.1016/j.crmeth.2021.100006
M3 - Article
AN - SCOPUS:85119343250
SN - 2667-2375
VL - 1
JO - Cell Reports Methods
JF - Cell Reports Methods
IS - 2
M1 - 100006
ER -