AUTHOR=Hurler Lisa , Toonen Erik J. M. , Kajdácsi Erika , van Bree Bregje , Brandwijk Ricardo J. M. G. E. , de Bruin Wieke , Lyons Paul A. , Bergamaschi Laura , Cambridge Institute of Therapeutic Immunology and Infectious Disease-National Institute of Health Research (CITIID-NIHR) COVID BioResource Collaboration , Sinkovits György , Cervenak László , Würzner Reinhard , Prohászka Zoltán , Baker Stephen , Bradley John R. , Chinnery Patrick F. , Cooper Daniel J. , Dougan Gordon , Goodfellow Ian G. , Gupta Ravindra K. , Kingston Nathalie , Lehner Paul J. , Lyons Paul A. , Matheson Nicholas J. , Saunders Caroline , Smith Kenneth G. C. , Summers Charlotte , Thaventhiran James , Torok M. Estee , Toshner Mark R. , Weekes Michael P. , Alvio Gisele , Baker Sharon , Bermperi Areti , Brookes Karen , Bucke Ashlea , Calder Jo , Canna Laura , Crucusio Cherry , Cruz Isabel , Jesus Rnalie de , Dempsey Katie , Stephano Giovanni Di , Domingo Jason , Elmer Anne , Harris Julie , Hewitt Sarah , Jones Heather , Jose Sherly , Kennet Jane , King Yvonne , Kourampa Jenny , Li Emily , McMahon Caroline , Meadows Anne , Mendoza Vivien , O’Brien Criona , Ocaya Charmain , Pascuale Ciro , Perales Marlyn , Price Jane , Rastall Rebecca , Ribeiro Carla , Rowlands Jane , Ruffolo Valentina , Tordesillas Hugo , Vargas Phoebe , Vergese Bensi , Watson Laura , Worsley Jieniean , Zerrudo Julie-Ann , Bergamaschi Laura , Betancourt Ariana , Bower Georgie , Bullman Ben , Cossetti Chiara , Sa Aloka De , Dunore Benjamin J. , Epping Maddie , Fawke Stuart , Gräf Stefan , Grenfell Richard , Hinch Andrew , Hodgson Josh , Huang Christopher , Huhn Oisin , Hunter Kelvin , Jarvis Isobel , Jones Emma , Josipović Maša , Legchenko Ekaterina , Lewis Daniel , Marsden Joe , Martin Jennifer , Mescia Federica , Nice Francesca , O’Donnell Ciara , Omarjee Ommar , Perera Marianne , Pointon Linda , Pond Nicole , Richoz Nathan , Romashova Nika , Savoinykh Natalia , Sharma Rahul , Shih Joy , Strezlecki Mateusz , Sutcliffe Rachel , Tilly Tobias , Tong Zhen , Treacy Carmen , Turner Lori , Wood Jennifer , Wylot Marta , Allison John , Biggs Heather , Butcher Helen , Caputo Daniela , Clapham-Riley Debbie , Dewhurst Eleanor , Fernandez Christian , Furlong Anita , Graves Barbara , Gray Jennifer , Ivers Tasmin , Gresley Emma Le , Linger Rachel , Kasanicki Mary , Meloy Sarah , Muldoon Francesca , Ovington Nigel , Papadia Sofia , Penkett Christopher J. , Phelan Isabel , Ranganath Venkatesh , Sambrook Jennifer , Schon Katherine , Stark Hannah , Stirrups Kathleen E. , Townsend Paul , Ziegenweidt Julie von , Webster Jennifer , Asaripour Ali , Mwaura Lucy , Patterson Caroline , Polwarth Gary , Bunclark Katherine , Mackay Michael , Michael Alice , Rossi Sabrina , Selvan Mayurun , Spencer Sarah , Yong Cissy , Polgarova Petra TITLE=Distinction of early complement classical and lectin pathway activation via quantification of C1s/C1-INH and MASP-1/C1-INH complexes using novel ELISAs JOURNAL=Frontiers in Immunology VOLUME=13 YEAR=2022 URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2022.1039765 DOI=10.3389/fimmu.2022.1039765 ISSN=1664-3224 ABSTRACT=

The most commonly used markers to assess complement activation are split products that are produced through activation of all three pathways and are located downstream of C3. In contrast, C4d derives from the cleavage of C4 and indicates either classical (CP) or lectin pathway (LP) activation. Although C4d is perfectly able to distinguish between CP/LP and alternative pathway (AP) activation, no well-established markers are available to differentiate between early CP and LP activation. Active enzymes of both pathways (C1s/C1r for the CP, MASP-1/MASP-2 for the LP) are regulated by C1 esterase inhibitor (C1-INH) through the formation of covalent complexes. Aim of this study was to develop validated immunoassays detecting C1s/C1-INH and MASP-1/C1-INH complex levels. Measurement of the complexes reveals information about the involvement of the respective pathways in complement-mediated diseases. Two sandwich ELISAs detecting C1s/C1-INH and MASP-1/C1-INH complex were developed and tested thoroughly, and it was investigated whether C1s/C1-INH and MASP-1/C1-INH complexes could serve as markers for either early CP or LP activation. In addition, a reference range for these complexes in healthy adults was defined, and the assays were clinically validated utilizing samples of 414 COVID-19 patients and 96 healthy controls. The immunoassays can reliably measure C1s/C1-INH and MASP-1/C1-INH complex concentrations in EDTA plasma from healthy and diseased individuals. Both complex levels are increased in serum when activated with zymosan, making them suitable markers for early classical and early lectin pathway activation. Furthermore, measurements of C1-INH complexes in 96 healthy adults showed normally distributed C1s/C1-INH complex levels with a physiological concentration of 1846 ± 1060 ng/mL (mean ± 2SD) and right-skewed distribution of MASP-1/C1-INH complex levels with a median concentration of 36.9 (13.18 - 87.89) ng/mL (2.5-97.5 percentile range), while levels of both complexes were increased in COVID-19 patients (p<0.0001). The newly developed assays measure C1-INH complex levels in an accurate way. C1s/C1-INH and MASP-1/C1-INH complexes are suitable markers to assess early classical and lectin pathway activation. An initial reference range was set and first studies showed that these markers have added value for investigating and unraveling complement activation in human disease.