Generation of a C3c specific monoclonal antibody and assessment of C3c as a putative inflammatory marker derived from complement factor C3

Yaseelan Palarasah; Karsten Skjodt; Jette Brandt; Børge Teisner; Claus Koch; Lars Vitved; Mikkel-Ole Skjoedt

doi:10.1016/j.jim.2010.09.024

Generation of a C3c specific monoclonal antibody and assessment of C3c as a putative inflammatory marker derived from complement factor C3

Yaseelan Palarasah, Karsten Skjodt, Jette Brandt, Børge Teisner, Claus Koch, Lars Vitved, Mikkel-Ole Skjoedt

18 Citations (Scopus)

Abstract

There is a general need for markers of systemic inflammation in acute or chronic diseases, where complement activation is involved. Available methods to monitor complement activation are elaborate and of low sensitivity; they include haemolytic assays (CH50), quantification of fluid phase terminal complex (C5b-C9) and quantification of complement split products by precipitation-in-gel techniques (e.g. C3d). We have developed a mouse monoclonal antibody (mAb) that is able to detect fluid phase C3c without interference from other products generated from the complement component C3. The C3c specific mAb was tested in different ELISA combinations with various types of in vitro activated sera and with plasma or serum samples from factor I deficient patients. The specificity of the mAb was evaluated in immunoprecipitation techniques and by analysis of eluted fragments of C3 after immunoaffinity chromatography. The C3c mAb was confirmed to be C3c specific, as it showed no cross-reactivity with native (un-cleaved) C3, with C3b, iC3b, or with C3d. Also, no significant reaction was observed with C3 fragments in factor I deficient sera or plasma. This antibody forms the basis for the generation of a robust ELISA that allows for a quick and reliable evaluation of complement activation and consumption as a marker for inflammatory processes. We established the C3c plasma range in 100 healthy Danish blood donors with a mean of 3.47 μg/ml and a range of 2.12-4.92 μg/ml. We believe that such an antibody might be of potential value in the assessment of in vivo complement activity during the inflammatory processes.

Original language	English
Journal	Journal of Immunological Methods
Volume	362
Issue number	1-2
Pages (from-to)	142-50
Number of pages	9
ISSN	0022-1759
DOIs	https://doi.org/10.1016/j.jim.2010.09.024
Publication status	Published - 31 Oct 2010

Keywords

Animals
Antibodies, Monoclonal
Antibody Specificity
Biomarkers
Blood Donors
Complement Activation
Complement C3c
Denmark
Enzyme-Linked Immunosorbent Assay
Female
Humans
Inflammation
Inflammation Mediators
Male
Mice
Mice, Inbred BALB C
Sensitivity and Specificity
Journal Article

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10.1016/j.jim.2010.09.024

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title = "Generation of a C3c specific monoclonal antibody and assessment of C3c as a putative inflammatory marker derived from complement factor C3",

abstract = "There is a general need for markers of systemic inflammation in acute or chronic diseases, where complement activation is involved. Available methods to monitor complement activation are elaborate and of low sensitivity; they include haemolytic assays (CH50), quantification of fluid phase terminal complex (C5b-C9) and quantification of complement split products by precipitation-in-gel techniques (e.g. C3d). We have developed a mouse monoclonal antibody (mAb) that is able to detect fluid phase C3c without interference from other products generated from the complement component C3. The C3c specific mAb was tested in different ELISA combinations with various types of in vitro activated sera and with plasma or serum samples from factor I deficient patients. The specificity of the mAb was evaluated in immunoprecipitation techniques and by analysis of eluted fragments of C3 after immunoaffinity chromatography. The C3c mAb was confirmed to be C3c specific, as it showed no cross-reactivity with native (un-cleaved) C3, with C3b, iC3b, or with C3d. Also, no significant reaction was observed with C3 fragments in factor I deficient sera or plasma. This antibody forms the basis for the generation of a robust ELISA that allows for a quick and reliable evaluation of complement activation and consumption as a marker for inflammatory processes. We established the C3c plasma range in 100 healthy Danish blood donors with a mean of 3.47 μg/ml and a range of 2.12-4.92 μg/ml. We believe that such an antibody might be of potential value in the assessment of in vivo complement activity during the inflammatory processes.",

keywords = "Animals, Antibodies, Monoclonal, Antibody Specificity, Biomarkers, Blood Donors, Complement Activation, Complement C3c, Denmark, Enzyme-Linked Immunosorbent Assay, Female, Humans, Inflammation, Inflammation Mediators, Male, Mice, Mice, Inbred BALB C, Sensitivity and Specificity, Journal Article",

author = "Yaseelan Palarasah and Karsten Skjodt and Jette Brandt and B{\o}rge Teisner and Claus Koch and Lars Vitved and Mikkel-Ole Skjoedt",

year = "2010",

month = oct,

day = "31",

doi = "10.1016/j.jim.2010.09.024",

language = "English",

volume = "362",

pages = "142--50",

journal = "Journal of Immunological Methods",

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TY - JOUR

T1 - Generation of a C3c specific monoclonal antibody and assessment of C3c as a putative inflammatory marker derived from complement factor C3

AU - Palarasah, Yaseelan

AU - Skjodt, Karsten

AU - Brandt, Jette

AU - Teisner, Børge

AU - Koch, Claus

AU - Vitved, Lars

AU - Skjoedt, Mikkel-Ole

PY - 2010/10/31

Y1 - 2010/10/31

N2 - There is a general need for markers of systemic inflammation in acute or chronic diseases, where complement activation is involved. Available methods to monitor complement activation are elaborate and of low sensitivity; they include haemolytic assays (CH50), quantification of fluid phase terminal complex (C5b-C9) and quantification of complement split products by precipitation-in-gel techniques (e.g. C3d). We have developed a mouse monoclonal antibody (mAb) that is able to detect fluid phase C3c without interference from other products generated from the complement component C3. The C3c specific mAb was tested in different ELISA combinations with various types of in vitro activated sera and with plasma or serum samples from factor I deficient patients. The specificity of the mAb was evaluated in immunoprecipitation techniques and by analysis of eluted fragments of C3 after immunoaffinity chromatography. The C3c mAb was confirmed to be C3c specific, as it showed no cross-reactivity with native (un-cleaved) C3, with C3b, iC3b, or with C3d. Also, no significant reaction was observed with C3 fragments in factor I deficient sera or plasma. This antibody forms the basis for the generation of a robust ELISA that allows for a quick and reliable evaluation of complement activation and consumption as a marker for inflammatory processes. We established the C3c plasma range in 100 healthy Danish blood donors with a mean of 3.47 μg/ml and a range of 2.12-4.92 μg/ml. We believe that such an antibody might be of potential value in the assessment of in vivo complement activity during the inflammatory processes.

AB - There is a general need for markers of systemic inflammation in acute or chronic diseases, where complement activation is involved. Available methods to monitor complement activation are elaborate and of low sensitivity; they include haemolytic assays (CH50), quantification of fluid phase terminal complex (C5b-C9) and quantification of complement split products by precipitation-in-gel techniques (e.g. C3d). We have developed a mouse monoclonal antibody (mAb) that is able to detect fluid phase C3c without interference from other products generated from the complement component C3. The C3c specific mAb was tested in different ELISA combinations with various types of in vitro activated sera and with plasma or serum samples from factor I deficient patients. The specificity of the mAb was evaluated in immunoprecipitation techniques and by analysis of eluted fragments of C3 after immunoaffinity chromatography. The C3c mAb was confirmed to be C3c specific, as it showed no cross-reactivity with native (un-cleaved) C3, with C3b, iC3b, or with C3d. Also, no significant reaction was observed with C3 fragments in factor I deficient sera or plasma. This antibody forms the basis for the generation of a robust ELISA that allows for a quick and reliable evaluation of complement activation and consumption as a marker for inflammatory processes. We established the C3c plasma range in 100 healthy Danish blood donors with a mean of 3.47 μg/ml and a range of 2.12-4.92 μg/ml. We believe that such an antibody might be of potential value in the assessment of in vivo complement activity during the inflammatory processes.

KW - Animals

KW - Antibodies, Monoclonal

KW - Antibody Specificity

KW - Biomarkers

KW - Blood Donors

KW - Complement Activation

KW - Complement C3c

KW - Denmark

KW - Enzyme-Linked Immunosorbent Assay

KW - Female

KW - Humans

KW - Inflammation

KW - Inflammation Mediators

KW - Male

KW - Mice

KW - Mice, Inbred BALB C

KW - Sensitivity and Specificity

KW - Journal Article

U2 - 10.1016/j.jim.2010.09.024

DO - 10.1016/j.jim.2010.09.024

M3 - Journal article

C2 - 20869965

SN - 0022-1759

VL - 362

SP - 142

EP - 150

JO - Journal of Immunological Methods

JF - Journal of Immunological Methods

IS - 1-2

ER -

Generation of a C3c specific monoclonal antibody and assessment of C3c as a putative inflammatory marker derived from complement factor C3

Abstract

Keywords

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