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complement components c1, c2, c3, c3b, c4 and factors h and i (CompTech Computer Technologies)

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CompTech Computer Technologies complement components c1, c2, c3, c3b, c4 and factors h and i

(A) Cofactor activities of allotypic variants of purified CFH (FH). In this graph of a representative assay, C3c release was measured (% cpm in supernatant) after 4 min at 37°C in cell-based assays of CFH cofactor activity with increasing concentrations of CFH (I62/Y402, V62/Y402 or V62/H402), factor I and 125I-EC3b cells. (B) Cleavage of <t>C3b</t> was measured in the fluid-phase assay using the I62/Y402, V62/Y402 or V62/H402 variants of purified CFH. Decay of the 115 kDa band (C3bα′) was determined by densitometry using ImageJ, expressed as a % of the 115 kDa band at time zero and plotted using SigmaPlot. (C) The rate constant for the exponential decay was measured using the equation y=a*exp(-b*x). There was no statistically significant difference in the rate constants between variants of CFH used. This held true over a range of CFH concentrations tested (5.5, 8.8 and 10.9 ng FH/μl) (p>0.07 for all comparisons of the different variants). [5.5 ng FH/μl], n=7; [8.8 ng FH/μl], n=3; and [10.9 ng FH/μl], n=5.

Complement Components C1, C2, C3, C3b, C4 And Factors H And I, supplied by CompTech Computer Technologies, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/complement+factors+c1%2C+c2%2C+c3+and+c4/pmc03639479-83-7-18?v=CompTech+Computer+Technologies
Average 90 stars, based on 1 article reviews

complement components c1, c2, c3, c3b, c4 and factors h and i - by Bioz Stars, 2026-07

90/100 stars

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1) Product Images from "Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration"

Article Title: Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration

Journal: Journal of immunology (Baltimore, Md. : 1950)

doi: 10.4049/jimmunol.0903596

Figure Legend Snippet: (A) Cofactor activities of allotypic variants of purified CFH (FH). In this graph of a representative assay, C3c release was measured (% cpm in supernatant) after 4 min at 37°C in cell-based assays of CFH cofactor activity with increasing concentrations of CFH (I62/Y402, V62/Y402 or V62/H402), factor I and 125I-EC3b cells. (B) Cleavage of C3b was measured in the fluid-phase assay using the I62/Y402, V62/Y402 or V62/H402 variants of purified CFH. Decay of the 115 kDa band (C3bα′) was determined by densitometry using ImageJ, expressed as a % of the 115 kDa band at time zero and plotted using SigmaPlot. (C) The rate constant for the exponential decay was measured using the equation y=a*exp(-b*x). There was no statistically significant difference in the rate constants between variants of CFH used. This held true over a range of CFH concentrations tested (5.5, 8.8 and 10.9 ng FH/μl) (p>0.07 for all comparisons of the different variants). [5.5 ng FH/μl], n=7; [8.8 ng FH/μl], n=3; and [10.9 ng FH/μl], n=5.

Techniques Used: Purification, Activity Assay

The rate of C3b cleavage is reduced by highly sulfated GAGs (heparin and over-sulfated HS) and accelerated by less N-sulfated GAGs (HS-BK and de-N-sulfated heparin). (A) Cleavage of C3b was measured in the fluid-phase assay using purified CFH with heparin (Hep) or HS-BK added. CFH (390 ng) and Factor I (200 ng) with or without 146 μg of a specific GAG were mixed and added to C3b (3.4 μg) in a total volume of 70 μl EDTA-VBS. At various time points, 10μl aliquots were removed and proteins separated on a gel as previously described. Decay of the 115 kDa band (C3bα′) was determined by densitometry using ImageJ, expressed as a % of the 115 kDa band at time zero and plotted using SigmaPlot. (B) Cleavage of C3b measured in the fluid-phase assay using purified CFH with sulfate-modified heparin (de-N- or de-O-sulfated) or over-sulfated HS added. (C) The rate of the fluid phase reaction is greatly reduced by over-sulfated HS (***, p< 0.001) and to a lesser extent heparin and de-O-sulfated heparin (**, p< 0.01) whereas HS-BK and de-N-sulfated heparin significantly increase the rate of this reaction (p< 0.001). Similar results were obtained using V62/Y402 or V62/H402 (data not shown). None (no added GAG), n=18; de-N-sulfated heparin, n=18; HS-BK, n=6; over-sulfated HS, n=9; de-O-sulfated heparin, n=6

Figure Legend Snippet: The rate of C3b cleavage is reduced by highly sulfated GAGs (heparin and over-sulfated HS) and accelerated by less N-sulfated GAGs (HS-BK and de-N-sulfated heparin). (A) Cleavage of C3b was measured in the fluid-phase assay using purified CFH with heparin (Hep) or HS-BK added. CFH (390 ng) and Factor I (200 ng) with or without 146 μg of a specific GAG were mixed and added to C3b (3.4 μg) in a total volume of 70 μl EDTA-VBS. At various time points, 10μl aliquots were removed and proteins separated on a gel as previously described. Decay of the 115 kDa band (C3bα′) was determined by densitometry using ImageJ, expressed as a % of the 115 kDa band at time zero and plotted using SigmaPlot. (B) Cleavage of C3b measured in the fluid-phase assay using purified CFH with sulfate-modified heparin (de-N- or de-O-sulfated) or over-sulfated HS added. (C) The rate of the fluid phase reaction is greatly reduced by over-sulfated HS (***, p< 0.001) and to a lesser extent heparin and de-O-sulfated heparin (**, p< 0.01) whereas HS-BK and de-N-sulfated heparin significantly increase the rate of this reaction (p< 0.001). Similar results were obtained using V62/Y402 or V62/H402 (data not shown). None (no added GAG), n=18; de-N-sulfated heparin, n=18; HS-BK, n=6; over-sulfated HS, n=9; de-O-sulfated heparin, n=6

Techniques Used: Purification, Modification

Figure Legend Snippet: Disaccharide analysis of Heparin and HSs used in C3b cleavage assays

Techniques Used:

Addition of human Bruch’s membrane/choroid punches increases the rate of cleavage of C3b by factor I and variants of CFH. (A) Cleavage of C3b was measured in the presence of four 6 mm diameter punches of Bruch’s membrane/choroid from the peripheral (two) and central (two) regions of posterior eyecups from a pair of human eyes (68-year-old donor) per reaction. Punches (four treated or four untreated with heparinase III) were added to CFH and factor I in ice cold EDTA-VBS. C3b was added, and the tubes placed at 37°C. The control tube had no punches added. At various times 18 μl was removed and run on a gel as previously described. Representative results from one isoform, I62/Y402, are depicted. (B) I62/Y402 and V62/H402 CFH both showed statistically significant increases in the rate constant in the presence of tissue (p< 0.01; n=5 each). There were no statistically significant differences between the variants of CFH alone compared to reactions with tissue pre-treated with heparanase III V62/H402). Data depicted are representative of 5 independent experiments from 5 pairs of donor eyes.

Figure Legend Snippet: Addition of human Bruch’s membrane/choroid punches increases the rate of cleavage of C3b by factor I and variants of CFH. (A) Cleavage of C3b was measured in the presence of four 6 mm diameter punches of Bruch’s membrane/choroid from the peripheral (two) and central (two) regions of posterior eyecups from a pair of human eyes (68-year-old donor) per reaction. Punches (four treated or four untreated with heparinase III) were added to CFH and factor I in ice cold EDTA-VBS. C3b was added, and the tubes placed at 37°C. The control tube had no punches added. At various times 18 μl was removed and run on a gel as previously described. Representative results from one isoform, I62/Y402, are depicted. (B) I62/Y402 and V62/H402 CFH both showed statistically significant increases in the rate constant in the presence of tissue (p< 0.01; n=5 each). There were no statistically significant differences between the variants of CFH alone compared to reactions with tissue pre-treated with heparanase III V62/H402). Data depicted are representative of 5 independent experiments from 5 pairs of donor eyes.

Techniques Used: Membrane, Control

Functional cofactor activity of CFH variant prebound to a single 6 mm Bruch’s membrane/choroid punch. (A) Cleavage of C3b measured in the fluid-phase assay using one Bruch’s membrane/choroid 6 mm punch preincubated with or without CFH (V62/Y402 or V62/H402) and washed, then added to VBS/EDTA, factor I and C3b. Aliquots were removed at 0, 2, 4, 8, 16, and 24 min and these samples were boiled and run with reducing buffer on 10% Bis-tris Criterion XT gels with MOPS buffer. After silver staining, the gel was scanned and the density of the 115 kDa band measured with ImageJ software, and SigmaPlot 9 used to create the graphs. The example shown is representative of 6 separate experiments. (B) Rate constants showed no significant differences between CFH variants independent of region tissue punch was taken from (i.e. periphery (as shown) or central) (p> 0.2).

Figure Legend Snippet: Functional cofactor activity of CFH variant prebound to a single 6 mm Bruch’s membrane/choroid punch. (A) Cleavage of C3b measured in the fluid-phase assay using one Bruch’s membrane/choroid 6 mm punch preincubated with or without CFH (V62/Y402 or V62/H402) and washed, then added to VBS/EDTA, factor I and C3b. Aliquots were removed at 0, 2, 4, 8, 16, and 24 min and these samples were boiled and run with reducing buffer on 10% Bis-tris Criterion XT gels with MOPS buffer. After silver staining, the gel was scanned and the density of the 115 kDa band measured with ImageJ software, and SigmaPlot 9 used to create the graphs. The example shown is representative of 6 separate experiments. (B) Rate constants showed no significant differences between CFH variants independent of region tissue punch was taken from (i.e. periphery (as shown) or central) (p> 0.2).

Techniques Used: Functional Assay, Activity Assay, Variant Assay, Membrane, Silver Staining, Software

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Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration

doi: 10.4049/jimmunol.0903596

Figure Lengend Snippet: (A) Cofactor activities of allotypic variants of purified CFH (FH). In this graph of a representative assay, C3c release was measured (% cpm in supernatant) after 4 min at 37°C in cell-based assays of CFH cofactor activity with increasing concentrations of CFH (I62/Y402, V62/Y402 or V62/H402), factor I and 125I-EC3b cells. (B) Cleavage of C3b was measured in the fluid-phase assay using the I62/Y402, V62/Y402 or V62/H402 variants of purified CFH. Decay of the 115 kDa band (C3bα′) was determined by densitometry using ImageJ, expressed as a % of the 115 kDa band at time zero and plotted using SigmaPlot. (C) The rate constant for the exponential decay was measured using the equation y=a*exp(-b*x). There was no statistically significant difference in the rate constants between variants of CFH used. This held true over a range of CFH concentrations tested (5.5, 8.8 and 10.9 ng FH/μl) (p>0.07 for all comparisons of the different variants). [5.5 ng FH/μl], n=7; [8.8 ng FH/μl], n=3; and [10.9 ng FH/μl], n=5.

Article Snippet: Human purified complement components C1, C2, C3, C3b, C4 and factors H and I were all obtained from CompTech, Inc (Tyler, TX).

Techniques: Purification, Activity Assay

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration

doi: 10.4049/jimmunol.0903596

Figure Lengend Snippet: The rate of C3b cleavage is reduced by highly sulfated GAGs (heparin and over-sulfated HS) and accelerated by less N-sulfated GAGs (HS-BK and de-N-sulfated heparin). (A) Cleavage of C3b was measured in the fluid-phase assay using purified CFH with heparin (Hep) or HS-BK added. CFH (390 ng) and Factor I (200 ng) with or without 146 μg of a specific GAG were mixed and added to C3b (3.4 μg) in a total volume of 70 μl EDTA-VBS. At various time points, 10μl aliquots were removed and proteins separated on a gel as previously described. Decay of the 115 kDa band (C3bα′) was determined by densitometry using ImageJ, expressed as a % of the 115 kDa band at time zero and plotted using SigmaPlot. (B) Cleavage of C3b measured in the fluid-phase assay using purified CFH with sulfate-modified heparin (de-N- or de-O-sulfated) or over-sulfated HS added. (C) The rate of the fluid phase reaction is greatly reduced by over-sulfated HS (***, p< 0.001) and to a lesser extent heparin and de-O-sulfated heparin (**, p< 0.01) whereas HS-BK and de-N-sulfated heparin significantly increase the rate of this reaction (p< 0.001). Similar results were obtained using V62/Y402 or V62/H402 (data not shown). None (no added GAG), n=18; de-N-sulfated heparin, n=18; HS-BK, n=6; over-sulfated HS, n=9; de-O-sulfated heparin, n=6

Article Snippet: Human purified complement components C1, C2, C3, C3b, C4 and factors H and I were all obtained from CompTech, Inc (Tyler, TX).

Techniques: Purification, Modification

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration

doi: 10.4049/jimmunol.0903596

Figure Lengend Snippet: Disaccharide analysis of Heparin and HSs used in C3b cleavage assays

Article Snippet: Human purified complement components C1, C2, C3, C3b, C4 and factors H and I were all obtained from CompTech, Inc (Tyler, TX).

Techniques:

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration

doi: 10.4049/jimmunol.0903596

Figure Lengend Snippet: Addition of human Bruch’s membrane/choroid punches increases the rate of cleavage of C3b by factor I and variants of CFH. (A) Cleavage of C3b was measured in the presence of four 6 mm diameter punches of Bruch’s membrane/choroid from the peripheral (two) and central (two) regions of posterior eyecups from a pair of human eyes (68-year-old donor) per reaction. Punches (four treated or four untreated with heparinase III) were added to CFH and factor I in ice cold EDTA-VBS. C3b was added, and the tubes placed at 37°C. The control tube had no punches added. At various times 18 μl was removed and run on a gel as previously described. Representative results from one isoform, I62/Y402, are depicted. (B) I62/Y402 and V62/H402 CFH both showed statistically significant increases in the rate constant in the presence of tissue (p< 0.01; n=5 each). There were no statistically significant differences between the variants of CFH alone compared to reactions with tissue pre-treated with heparanase III V62/H402). Data depicted are representative of 5 independent experiments from 5 pairs of donor eyes.

Article Snippet: Human purified complement components C1, C2, C3, C3b, C4 and factors H and I were all obtained from CompTech, Inc (Tyler, TX).

Techniques: Membrane, Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Heparan Sulfate, Including that in Bruch's Membrane, Inhibits the Complement Alternative Pathway: Implications for Age-related Macular Degeneration

doi: 10.4049/jimmunol.0903596

Figure Lengend Snippet: Functional cofactor activity of CFH variant prebound to a single 6 mm Bruch’s membrane/choroid punch. (A) Cleavage of C3b measured in the fluid-phase assay using one Bruch’s membrane/choroid 6 mm punch preincubated with or without CFH (V62/Y402 or V62/H402) and washed, then added to VBS/EDTA, factor I and C3b. Aliquots were removed at 0, 2, 4, 8, 16, and 24 min and these samples were boiled and run with reducing buffer on 10% Bis-tris Criterion XT gels with MOPS buffer. After silver staining, the gel was scanned and the density of the 115 kDa band measured with ImageJ software, and SigmaPlot 9 used to create the graphs. The example shown is representative of 6 separate experiments. (B) Rate constants showed no significant differences between CFH variants independent of region tissue punch was taken from (i.e. periphery (as shown) or central) (p> 0.2).

Article Snippet: Human purified complement components C1, C2, C3, C3b, C4 and factors H and I were all obtained from CompTech, Inc (Tyler, TX).

Techniques: Functional Assay, Activity Assay, Variant Assay, Membrane, Silver Staining, Software

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