Levels of mannan-binding lectin-associated serine protease-2 in healthy individuals Mette Møller-Kristensen a, * , Jens Chr. Jensenius a , Lisbeth Jensen a , Nicole Thielens b , Ve ´ronique Rossi b , Gerard Arlaud b , Steffen Thiel a a Department of Medical Microbiology and Immunology, University of Aarhus, Aarhus C 8000, Denmark b Laboratoire d’Enzymologie Mole ´culaire, Institut de Biologie Structurale Jean-Pierre Ebel, Grenoble, France Received 24 June 2003; received in revised form 19 August 2003; accepted 21 August 2003 Abstract The lectin pathway is part of the innate immune system providing a first line of defence against infections. Mannan-binding lectin (MBL) and ficolins, in complex with MBL-associated serine proteases (MASPs), are capable of activating the complement system, thus mediating the destruction of infectious agents. MASP-2 cleaves C4 and C2 and is thus crucial for the activation of downstream complement components. We present an assay for quantifying total MASP-2 in plasma and serum samples. The assay is a sandwich type assay using a combination of two monoclonal anti-MASP-2 antibodies, one directed against the N-terminal part of MASP-2 and the other against its C-terminal part. Based on a population of Danish blood donors, the average MASP-2 concentration was estimated at 534 (S.D. F 213) ng per ml of plasma. Characterization of the MASP-2 protein in serum showed high stability at 4 jC and at ambient temperature but a rapid decline at 37 jC. Gel permeation chromatography (GPC) indicated that all MASP-2 in serum is present in complexes with MBL and ficolins. D 2003 Elsevier B.V. All rights reserved. Keywords: Mannan-binding lectin-associated serine proteases; MASP; Mannose-binding lectin; Ficolin; TRIFMA 1. Introduction The ability of lectins, i.e., carbohydrate-binding proteins, and other pattern recognition molecules to distinguish between self and non-self is one of the characteristics of innate immunity. Mannan-binding lectin (MBL) and L- and H-ficolin detect pathogen- associated molecular patterns (PAMPs) and promote destruction of pathogens by activating effector mech- anisms of the complement system (Holmskov et al., 2003). MBL shows selectivity for carbohydrates pre- senting 3- and 4-OH groups in the equatorial plane of the ring structure (Weis et al., 1992), while the epitopes recognized by ficolins have not been defined, but are present in, e.g., N-acetyl-glucosamine (GlcNAc) (Matsushita et al., 1996). For activating the complement system, MBL, L- and H-ficolin rely on associated serine proteases. These were first discovered in complex with MBL and are referred to as MBL-associated serine proteases (MASPs). MBL, L- and H-ficolin (Matsushita et al., 2000a, 2002) are associated with three MASPs, termed MASP-1 (Matsushita and Fujita, 1992), MASP-2 0022-1759/$ - see front matter D 2003 Elsevier B.V. All rights reserved. doi:10.1016/j.jim.2003.08.012 * Corresponding author. Tel.: +45-89-421778; fax: +45-86- 196128. E-mail address: mmk@microbiology.au.dk (M. Møller-Kristensen). www.elsevier.com/locate/jim Journal of Immunological Methods 282 (2003) 159 – 167