Details
| Original language | English |
|---|---|
| Pages (from-to) | 3368-3371 |
| Number of pages | 4 |
| Journal | Analytical chemistry |
| Volume | 65 |
| Issue number | 23 |
| Publication status | Published - 1 Dec 1993 |
| Externally published | Yes |
Abstract
A sandwich-type flow-injection binding assay for quantitation of various IgG's was developed. The assay is based on the pseudoimmunological reaction between protein A from Staphylococcus aureus and immunoglobulin G from different species. Protein A immobilized on a solid support and a fusion protein of protein A and β-galactosidase from Escherichia coli are used for detection. The fusion protein is produced with a temperature-inducible recombinant E. coli strain. A sandwich is formed by subsequent injection of IgG and fusion protein into the buffer stream flowing through the immobilized protein A column. The amount of enzyme activity bound is proportional to the amount of IgG bound and is measured by pumping a lactose solution as substrate for β-galactosidase through the protein A column. Lactose is converted to glucose and galactose. The detector is an enzyme thermistor that measures the heat evolved in the enzymatic conversion of glucose by coimmobilized glucose oxidase and catalase. The assay takes 16 min at a flow rate of 0.6 mL min-1 with a lower detection limit of 33 pmol per injection of rabbit IgG. The precision of replicate measurements has a standard deviation of 4–5%, and the column can be used for more than 50 cycles.
ASJC Scopus subject areas
- Chemistry(all)
- Analytical Chemistry
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In: Analytical chemistry, Vol. 65, No. 23, 01.12.1993, p. 3368-3371.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Specific Flow Injection Sandwich Binding Assay for IgG Using Protein A and a Fusion Protein
AU - Brandes, Wiebke
AU - Maschke, Hans Eckhard
AU - Scheper, Thomas
PY - 1993/12/1
Y1 - 1993/12/1
N2 - A sandwich-type flow-injection binding assay for quantitation of various IgG's was developed. The assay is based on the pseudoimmunological reaction between protein A from Staphylococcus aureus and immunoglobulin G from different species. Protein A immobilized on a solid support and a fusion protein of protein A and β-galactosidase from Escherichia coli are used for detection. The fusion protein is produced with a temperature-inducible recombinant E. coli strain. A sandwich is formed by subsequent injection of IgG and fusion protein into the buffer stream flowing through the immobilized protein A column. The amount of enzyme activity bound is proportional to the amount of IgG bound and is measured by pumping a lactose solution as substrate for β-galactosidase through the protein A column. Lactose is converted to glucose and galactose. The detector is an enzyme thermistor that measures the heat evolved in the enzymatic conversion of glucose by coimmobilized glucose oxidase and catalase. The assay takes 16 min at a flow rate of 0.6 mL min-1 with a lower detection limit of 33 pmol per injection of rabbit IgG. The precision of replicate measurements has a standard deviation of 4–5%, and the column can be used for more than 50 cycles.
AB - A sandwich-type flow-injection binding assay for quantitation of various IgG's was developed. The assay is based on the pseudoimmunological reaction between protein A from Staphylococcus aureus and immunoglobulin G from different species. Protein A immobilized on a solid support and a fusion protein of protein A and β-galactosidase from Escherichia coli are used for detection. The fusion protein is produced with a temperature-inducible recombinant E. coli strain. A sandwich is formed by subsequent injection of IgG and fusion protein into the buffer stream flowing through the immobilized protein A column. The amount of enzyme activity bound is proportional to the amount of IgG bound and is measured by pumping a lactose solution as substrate for β-galactosidase through the protein A column. Lactose is converted to glucose and galactose. The detector is an enzyme thermistor that measures the heat evolved in the enzymatic conversion of glucose by coimmobilized glucose oxidase and catalase. The assay takes 16 min at a flow rate of 0.6 mL min-1 with a lower detection limit of 33 pmol per injection of rabbit IgG. The precision of replicate measurements has a standard deviation of 4–5%, and the column can be used for more than 50 cycles.
UR - http://www.scopus.com/inward/record.url?scp=0027746255&partnerID=8YFLogxK
U2 - 10.1021/ac00071a006
DO - 10.1021/ac00071a006
M3 - Article
C2 - 8297025
AN - SCOPUS:0027746255
VL - 65
SP - 3368
EP - 3371
JO - Analytical chemistry
JF - Analytical chemistry
SN - 0003-2700
IS - 23
ER -