Biotinylation of human C3

Berger, Merle J.; Cole, Roger M.; Gaither, J.A.; Brown, E M; Chused, Thomas M.; Hammer, Carl H.; Frank, M.M.

doi:10.1016/0161-5890(82)90018-9

Cited by 5 publications

(7 citation statements)

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“…OXI005 antibody was biotinylated according to standard procedures essentially as described 13 using Biotin-X-NHS (Merck, Darmstadt, Germany) in a phosphate-buffered saline (PBS) buffer at pH 7.2.…”

Section: Biotinylated Antibodymentioning

confidence: 99%

A Luminescent Oxygen Channeling Immunoassay for the Determination of Insulin in Human Plasma

Poulsen¹,

Jensen²

2007

SLAS Discovery

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The authors describe a homogeneous, sensitive, and rapid bead-based sandwich immunoassay with a broad analytical range for quantifying insulin in human plasma. The assay was performed as a 2-step reaction by incubating the sample with a mixture of biotinylated anti-insulin antibody and beads covalently coated with anti-insulin antibody for 1 h. This was followed by incubation with beads covalently coated with streptavidin for 30 min. After the incubation steps, light generated from a chemiluminescent reaction within the beads was quantitated. The assay was run in 384-well plates with a sample volume of 5 microL. The analytical range extended from 1 to 10,000 pM. Intra-assay precision (% coefficient of variation) ranged from 1.9% to 3.8% for various insulin concentrations. Interassay precision ranged from 4.6% to 7.3%. Assay detection limit was 0.3 pM. There was no interference from moderate hemolysis (with hemoglobin up to 375 mg/dL), bilirubin (up to at least 50 mg/dL), triglyceride (up to at least 1000 mg/dL), biotin (up to at least 7.7 ng/mL), or ascorbic acid (up to 100 mg/dL). However, gross hemolysis did affect the assay. Comparable results were obtained for plasma (ethylenediamine tetra-acetic acid, citrate, and heparin treated) and serum. The correlation with enzyme-linked immunosorbent assay (ELISA) was good (y = 1.25x + 1.19, R(2) = 0.98). This method is convenient and represents an alternative to ELISA.

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Section: Biotinylated Antibodymentioning

confidence: 99%

A Luminescent Oxygen Channeling Immunoassay for the Determination of Insulin in Human Plasma

Poulsen¹,

Jensen²

2007

SLAS Discovery

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“…Hopkins and colleagues (1979a, b; Tolson et al, 1981) used avidin-stabilized gold colloids as markers for cell surface proteins on granulosa cells. In addition to these reports, others have used the avidin-biotin system to visually demonstrate human C3 (Berger et al, 1982) on erythrocytes and neural antigens (Van Den Pol, 1984). Thus, the avidin-biotin system has shown a wide variety of applications.…”

Section: Discussionmentioning

confidence: 99%

Problems in the production and use of 5 nm avidin‐gold colloids

Morris¹,

Saelinger²

1986

Journal of Microscopy

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SUMMARY Over the past 5 years we have encountered several problems in the production and use of 5 nm avidin‐gold colloids for markers in electron microscopy. These problems include flocculation of colloids during reduction of chloroauric acid, insoluble gold pellets following ultracentrifugation, and non‐specific binding of avidin‐gold colloids to biological membranes. We are able to avoid these problems by: (1) avoiding the use of crystalline chloroauric acid; (2) succinoylating egg white avidin prior to adsorption on the gold sols; (3) resuspending the pellets following ultracentrifugation in 5 mM phosphate buffer, pH 7.5; and (4) using the avidin‐gold colloids within 4 weeks of production.

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“…Guinea pig Cl was purchased from Cordis Laboratories, Miami, Fla. Functionally pure human C4 and C2 were prepared as previously described (5). Biochemically pure C3 was prepared and biotinylated as previously described (2). In some experiments, C3 and biotinyl-C3 were radiolabeled with 1251 via the Bolton-Hunter reagent, as previously described (3), to a specific activity of 105 cpm/p.g without loss of hemolytic activity.…”

Section: Methodsmentioning

confidence: 99%

Localization of complement component 3 on Streptococcus pneumoniae: anti-capsular antibody causes complement deposition on the pneumococcal capsule

Brown¹,

Joiner²,

Cole³

et al. 1983

Infect Immun

101

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We have previously shown that complement component 3 (C3) deposited onto encapsulated Streptoc-occius pnzeiotuttiiae by anti-capsular antibody (Ab) is a more efficient opsonin in vitro and in vivo than C3 deposited by anti-cell wall Ab (Brown et al., J. Clin. Invest. 69:85-98, 1982). In the present study, we explored the cellular location of C3b molecules that differ in opsonic efficiency by using avidin-ferritin to localize biotinylated Ab and C3 molecules on S. pnleiuitiolniae for electron microscopy. Anti-cell wall Ab and C3b molecules deposited by this Ab on unencapsulated S. pneimsoniae were localized to S. pnteiottiae cell walls. Anti-capsular Ab and C3b deposited by this Ab were seen in clusters on encapsulated S. pneuinoniiae at a distance from the cell wall. However, no avidinferritin staining of encapsulated S. pneitu,notn(ie was seen on incubation with biotinyl-anti-cell wall Ab. biotinylated C3 fixed by anti-cell wall Ab, or nonimmune serum containing biotinyl-C3. In each case, uptake of the biotinylated component was proven by radioactivity measurements, since biotinylated Ab and

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Biotinylation of human C3

Cited by 5 publications

References 0 publications

A Luminescent Oxygen Channeling Immunoassay for the Determination of Insulin in Human Plasma

A Luminescent Oxygen Channeling Immunoassay for the Determination of Insulin in Human Plasma

Problems in the production and use of 5 nm avidin‐gold colloids

Localization of complement component 3 on Streptococcus pneumoniae: anti-capsular antibody causes complement deposition on the pneumococcal capsule

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