2021
DOI: 10.1002/eji.202149319
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Single‐reaction multi‐antigen serological test for comprehensive evaluation of SARS‐CoV‐2 patients by flow cytometry

Abstract: Here, we describe a new, simple, highly multiplexed serological test that generates a more complete picture of seroconversion than single antigen‐based assays. Flow cytometry is used to detect multiple Ig isotypes binding to four SARS‐CoV‐2 antigens: the Spike glycoprotein, its RBD fragment (the main target for neutralizing antibodies), the nucleocapsid protein, and the main cysteine‐like protease in a single reaction. Until now, most diagnostic serological tests measured antibodies to only one antigen and in … Show more

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Cited by 11 publications
(6 citation statements)
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“…Higher epitope density likely allows for monogamous multivalent binding of low affinity antibodies. Other immunoassays to detect specific anti-Spike antibodies by flow cytometry have been developed and either do not show IgM detection ( 20 , 21 ) or they do efficiently detect IgM on beads where the antigen has been adsorbed ( 22 24 ) or on the surface of HEK-293T cells where the antigen is overexpressed ( 25 28 ). In these two approaches epitope density is higher compared to the Jurkat-Spike cells of the JFCI, in line with our argument.…”
Section: Discussionmentioning
confidence: 99%
“…Higher epitope density likely allows for monogamous multivalent binding of low affinity antibodies. Other immunoassays to detect specific anti-Spike antibodies by flow cytometry have been developed and either do not show IgM detection ( 20 , 21 ) or they do efficiently detect IgM on beads where the antigen has been adsorbed ( 22 24 ) or on the surface of HEK-293T cells where the antigen is overexpressed ( 25 28 ). In these two approaches epitope density is higher compared to the Jurkat-Spike cells of the JFCI, in line with our argument.…”
Section: Discussionmentioning
confidence: 99%
“…Purified Spike protein was prepared as described in [ 20 ]. Briefly, a recombinant cDNA cloned in pCDNA3.1 vector and coding for soluble S protein containing a T4 fibritin trimerization sequence, a Flag epitope and an 8xHistag (all at the C-terminus), was transfected in HEK293F cells using standard procedures.…”
Section: Methodsmentioning
confidence: 99%
“…To the best of our knowledge, there is only three studies employing a cell‐free flow cytometric approach to evaluate the humoral immune response (Cáceres‐Martell et al . 2021 ; Dogan et al . 2021 ; Egia‐Mendikute et al .…”
Section: Resultsmentioning
confidence: 99%
“…These beads were used for simultaneous detection of IgG, IgM and IgA by flow cytometry, showing results with low background signals and high specificity and sensitivity (Cáceres‐Martell et al . 2021 ). However, magnetic beads are not compatible with certain flow cytometers.…”
Section: Resultsmentioning
confidence: 99%
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