Monitoring of dynamic changes in Keyhole Limpet Hemocyanin (KLH)-specific B cells in KLH-vaccinated cancer patients.
Keyhole limpet hemocyanin (KLH) is used as an immunogenic neo-antigen for various clinical applications and during vaccine development. For advanced monitoring of KLH-based interventions, we developed a flow cytometry-based assay for the ex vivo detection, phenotyping and isolation of KLH-specific B cells. As proof-of-principle, we analyzed 10 melanoma patients exposed to KLH during anti-cancer dendritic cell vaccination. Our assay demonstrated sensitive and specific detection of KLH-specific B cells in peripheral blood and KLH-specific B cell frequencies strongly correlated with anti-KLH serum antibody titers. Profiling of B cell subsets over the vaccination course revealed that KLH-specific B cells matured from naïve to class-switched memory B cells, confirming the prototypic B cell response to a neo-antigen. We conclude that flow-cytometric detection and in-depth phenotyping of KLH-specific B cells is specific, sensitive, and scalable. Our findings provide novel opportunities to monitor KLH-specific immune responses and serve as a blueprint for the development of new flow-cytometric protocols.
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud university medical center, Nijmegen, The Netherlands.
Florian
F
Wimmers
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud university medical center, Nijmegen, The Netherlands.
Nienke
N
de Haas
Department of Medical Microbiology, Radboud university medical center, Nijmegen, The Netherlands.
Anja
A
Scholzen
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud university medical center, Nijmegen, The Netherlands.
Gerty
G
Schreibelt
Department of Pediatrics, Laboratory of Pediatric Infectious Diseases, Radboud university medical center, Nijmegen, The Netherlands.
Elles
E
Simonetti
Department of Pediatrics, Laboratory of Pediatric Infectious Diseases, Radboud university medical center, Nijmegen, The Netherlands.
Marc J
MJ
Eleveld
Department of Laboratory Medicine, Laboratory Medical Immunology, Radboud university medical center, Nijmegen, The Netherlands.
Huberdina M L M
HM
Brouwers
Department of Laboratory Medicine, Laboratory Medical Immunology, Radboud university medical center, Nijmegen, The Netherlands.
Marjo
M
Beldhuis-Valkis
Department of Laboratory Medicine, Laboratory Medical Immunology, Radboud university medical center, Nijmegen, The Netherlands.
Irma
I
Joosten
Department of Pediatrics, Laboratory of Pediatric Infectious Diseases, Radboud university medical center, Nijmegen, The Netherlands.
Marien I
MI
de Jonge
Department of Medical Oncology, Radboud university medical center, Nijmegen, The Netherlands.
Winald R
WR
Gerritsen
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud university medical center, Nijmegen, The Netherlands.|Department of Medical Oncology, Radboud university medical center, Nijmegen, The Netherlands.
I Jolanda M
IJ
de Vries
Department of Pediatrics, Laboratory of Pediatric Infectious Diseases, Radboud university medical center, Nijmegen, The Netherlands.
Dimitri A
DA
Diavatopoulos
Department of Laboratory Medicine, Laboratory Medical Immunology, Radboud university medical center, Nijmegen, The Netherlands.
Joannes F M
JF
Jacobs
D000276:Adjuvants, Immunologic / Q000737:chemistry*; D000906:Antibodies / Q000097:blood; D016175:B-Lymphocyte Subsets / Q000145:classification / Q000276:immunology* / Q000473:pathology; D019496:Cancer Vaccines / Q000627:therapeutic use*; D058948:Cell Tracking / Q000379:methods; D003713:Dendritic Cells / Q000737:chemistry / Q000166:cytology / Q000276:immunology / Q000637:transplantation*; D058501:Enzyme-Linked Immunospot Assay; D005434:Flow Cytometry; D006433:Hemocyanins / Q000737:chemistry*; D006801:Humans; D007156:Immunologic Memory; D016130:Immunophenotyping; D008545:Melanoma / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D012878:Skin Neoplasms / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D014611:Vaccination / Q000379:methods
D000276:Adjuvants, Immunologic; D000906:Antibodies; D019496:Cancer Vaccines; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin