Computer-Assisted Kinetic Assay for Quantification of Total Complement Activity

Objective. To develop a novel sensitive and accurate assay suitable for high-volume testing of the total complement activity in the serum for clinical laboratories. Methods. The total complement activity (TCA) to be measured was quantified by detecting the number of fragments produced by erythrocyte lysis and the erythrocyte fragmentation index (EFI), indicating TCA. EFI = M × M 2 / M 1 + M 2 , where M is the number of erythrocyte fragments (removed from the background), M 1 is the number of unagglutinated red cells, M 2 is the number of agglutinated red cell groups, and M 2 / M 1 + M 2 is the agglutination coefficient indicating the degree of erythrocyte agglutination. Mild changes in hemolysin and erythrocyte concentrations were made to optimize the testing conditions. The same serum samples were tested for 10 consecutive days to determine the stability of the experimental results. Serum EFI was detected in both nephrotic syndrome patients and healthy subjects. Results. There was a linear relationship between hemolysin and erythrocyte agglutination ( r = 0.999 , P < 0.001 ). A good linear relationship existed between EFI and TCA ( r = 0.991 , P < 0.001 ). The results were not affected by slight fluctuations in the concentrations of hemolysin or erythrocytes. The interbatch CV = 8.6 % of the test results showed good stability. There was a significant difference in the EFI between nephrotic syndrome patients and healthy individuals, P < 0.001 , and EFI was reduced in nephrotic syndrome patients compared to healthy individuals. Conclusion. The flow cytometry-based assay for TCA was sensitive and accurate and had potential value for clinical application.

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THE DEVELOPMENT OF TOTAL COMPLEMENT ACTIVITY ASSAY BASED ON COMPLEMENT-DEPENDENT IMMUNE LIPOSOME LYSIS

Annals of the Russian academy of medical sciences ◽

10.15690/vramn.v69.i3-4.991 ◽

2015 ◽

Vol 69 (3-4) ◽

pp. 24-30

Author(s):

S. N. Skopinskaya ◽

S. P. Yarkov ◽

E. N. Khramov ◽

A. V. Antashev

Keyword(s):

Blood Serum ◽

Activity Assay ◽

Animal Blood ◽

Complement Activity ◽

Sheep Erythrocytes ◽

Wide Range ◽

Total Complement ◽

Sulforhodamine 101 ◽

Internal Volume ◽

Definition Of

Background: The purpose of work was development of a fast and reproduced procedure for measurement of the total complement activity (ТСА) in human or animal blood serum. Materials and methods: Steady at storage liposomes preparations, which surface sensitized 2,4-DNP haptens, and the internal volume contains calceine or sulforhodamine 101 are obtained. Complement-dependent immune lysis of liposomes at presence of the anti-2,4-DNP immunoglobulines and complement preparations from animals are investigated. Results: It is shown that the degree of liposomes immune lysis depends on complement concentration in a wide range that can be used for definition of TCA level. Research of blood sera from patients has revealed correlation (r =0,793) between data received with the help of liposome immunolytic systems, and the data of nephelometric analysis with application of suspension sheep erythrocytes. Conclusion: The method allows to define total complement activity in blood serum in 15 minutes without separation of reaction components. This might be useful for measurement ТСА level at patients with various diseases and realization of scientific researches.

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Evaluation of the Optilite® analyser for determination of total complement activity and C3 and C4 fractions

Annales de biologie clinique ◽

10.1684/abc.2019.1453 ◽

2019 ◽

Vol 77 (4) ◽

pp. 447-452

Author(s):

Benoit Nespola ◽

Hélène Comitogianni ◽

Isabelle Jahn ◽

Joëlle Goetz

Keyword(s):

Complement Activity ◽

Total Complement

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Automated homogeneous liposome-based assay system for total complement activity

Clinical Chemistry ◽

10.1093/clinchem/41.4.586 ◽

1995 ◽

Vol 41 (4) ◽

pp. 586-590 ◽

Cited By ~ 25

Author(s):

S Yamamoto ◽

K Kubotsu ◽

M Kida ◽

K Kondo ◽

S Matsuura ◽

...

Keyword(s):

Serum Proteins ◽

M Protein ◽

Homogeneous Population ◽

Complement Activity ◽

Activity Test ◽

Human Sera ◽

Total Complement ◽

Hemolytic Complement Activity ◽

Glucose 6 Phosphate Dehydrogenase

Abstract We developed an automated homogeneous immunoassay, based on immune lysis of dinitrophenyl (DNP)-labeled liposomes, for measuring total complement activity. Liposome lysis caused by complement activity was detected spectrophotometrically from entrapped glucose-6-phosphate dehydrogenase activity. Complement activity in human sera was quantified by comparison with a calibration curve. For ease of application to fully automated routine clinical analyzers, we adopted a two-reagent system, one reagent containing a homogeneous population of small DNP-labeled liposomes and one containing antibody/substrate. This system required calibration only once a week. Within-run and between-run CVs were 0.4-1.3% (n = 10) and 1.8-4.7% (n = 10), respectively. Serum results were linear upon dilution (with saline) over a twofold range. Bilirubin, hemoglobin, Intrafat, and serum proteins such as rheumatoid factor, M protein, IgG, and IgA did not affect the assay results. The results (y) correlated well with those from a hemolytic complement activity test (x): y = 1.05x - 1.14, r = 0.92, on 66 samples in the range < 10- > 50 kU/L. This method should therefore be of great use for the determination of complement activity.

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Establishment of a method for measuring total complement activity based on a hemolysis system using own red blood cells

Journal of Immunological Methods ◽

10.1016/j.jim.2016.01.010 ◽

2016 ◽

Vol 430 ◽

pp. 21-27 ◽

Cited By ~ 1

Author(s):

Ruirui Dong ◽

Hui Liu

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Complement Activity ◽

Total Complement

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Differences of Total Activity and Heat Tolerance of Complement in Plasma Between Black and Yellow Populations

10.21203/rs.2.19294/v1 ◽

2019 ◽

Author(s):

Abdelhakam G. Tamomh ◽

Xiaojun Jin ◽

Hui Liu

Keyword(s):

Heat Tolerance ◽

Total Activity ◽

Ratio Method ◽

Healthy Individuals ◽

Blood Samples ◽

Complement Activity ◽

Temperature Calculation ◽

Total Complement ◽

Standard Reaction ◽

Tolerance Distribution

Abstract Background The goal of this paper is to compare and evaluate the differences in total activity and Heat Tolerance of complement distribution Between Black and Yellow populations.Methods Blood samples of Black and Yellow healthy individuals were randomly collected, and plasma was obtained. The test plasma was diluted in a five series concentration, following with standard reaction and experimental reaction measurements. The basic principle of heat tolerance temperature calculation is to consider the ratio value (ratio method) in which heat tolerance of TCA calculated and plotted according to the area of trapezoidal and triangle as follows: C x°C=OD1-OD5+(OD2-OD5+OD3-OD5+OD4-OD5)*2. Where Cx°C is the total complement activity of the specimen measured in the two reaction conditions.Results The total activity of complement in Black individuals was statistically significantly lower than Yellow individuals (P< 0.05). The heat tolerance of total complement activity was higher among blacks than yellow individuals with no differences between the two groups (P> 0.05). The frequency values of total activity of complement distributed in the yellow individuals was statistically higher than Blacks (P<0.05) and The heat tolerance of total complement activity was statistically higher significant among Blacks than Yellow individuals (P<0.05).Conclusions There is a high differences in total complement activity and heat Tolerance distribution between Black and Yellow populations, may predict health status and innate immunity level differences between African race and Asian race.

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Homogeneous, liposome-based assay for total complement activity in serum.

Clinical Chemistry ◽

10.1093/clinchem/32.2.275 ◽

1986 ◽

Vol 32 (2) ◽

pp. 275-278 ◽

Cited By ~ 11

Author(s):

D W Bowden ◽

M Rising ◽

G Akots ◽

A Myles ◽

R J Broeze

Keyword(s):

Normal Population ◽

Test Results ◽

Normal Range ◽

Serum Samples ◽

Complement Components ◽

Complement Activity ◽

Standard Curve ◽

Total Complement ◽

Speed Accuracy ◽

Range Test

Abstract This is a rapid, homogeneous, liposome-based assay for total complement activity in human serum. Liposome-encapsulated enzyme is unmasked by the action of complement on liposomes carrying surface-bound immune complexes. The amount of unmasked enzyme, proportional to the concentration of added complement, is quantified by measuring the absorbance of enzymically produced product at 410 nm. Complement activity in serum samples is extrapolated from a standard curve generated from dilutions of a guinea pig serum containing a known activity of complement. Interassay CVs were less than 7.0% and intra-assay CVs less than 2.8% for serum pools with complement activities spanning the normal range. Test results correlate as well with those of the hemolytic complement test (r = 0.80) as the latter correlates with itself (r = 0.82), and also correlate reasonably with measurements of complement components C3 (r = 0.62) and C4 (r = 0.74). Values for a normal population are reported. Advantages of this test include stability of reagents, speed, accuracy, simplicity, and avoidance of radioisotopes.

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