Heterogeneous enzyme immunoassay of alpha-fetoprotein in maternal serum by flow-injection amperometric detection of 4-aminophenol

1990 ◽  
Vol 36 (11) ◽  
pp. 1941-1944 ◽  
Author(s):  
Y Xu ◽  
B Halsall ◽  
W R Heineman
Keyword(s):  
Enzyme Immunoassay ◽  
Flow Injection ◽  
Enzymatic Reaction ◽  
Amperometric Detection ◽  
Maternal Serum ◽  
Alpha Fetoprotein ◽  
Serum Samples ◽  
Low Concentrations ◽  
Sandwich Type ◽  
Commercial Kit

Abstract A sandwich-type heterogeneous enzyme immunoassay with flow-injection analysis for alpha-fetoprotein (AFP) in human serum has been developed. 4-Aminophenol, the product of enzymatic reaction, is detected amperometrically. The interassay CV for this electrochemical enzyme immunoassay was less than 8.2%, with a minimum detection limit for AFP of 0.163 micrograms/L. The calibration curve had a linear range of 0.316-100 micrograms/L. Studies with 48 human maternal serum samples, comparing results by this method with those by a commercial kit, showed a good correlation (r = 0.961). This procedure provides an alternative method for determining low concentrations of AFP in human maternal serum.

Specific enzyme immunoassay for human chorionic gonadotrophin

1981 ◽  
Vol 97 (4) ◽  
pp. 562-568 ◽  
Author(s):  
Tatsuhiro Sekiya ◽  
Yoshihito Furuhashi ◽  
Setsuko Goto ◽  
Shigeaki Kaseki ◽  
Yutaka Tomoda ◽  
...  
Keyword(s):  
Enzyme Immunoassay ◽  
Solid Phase ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Serum Samples ◽  
Coefficients Of Variation ◽  
Highly Sensitive ◽  
Assay Tube ◽  
Sandwich Type ◽  
Serial Samples

Abstract. A sandwich-type enzyme immunoassay system specific for human chorionic gonadotrophin (hCG) was prepared with the antibody Fab'-β-D-galactosidase complex and the antibody F(ab')2-immobilized silicone rubber solid phase by using a purified antibody to β subunit of hCG (hCGβ). The assay system cross-reacted less than 4% with human luteinizing hormone (hLH) and human follicular stimulating hormone (hFSH), and proved to be highly sensitive with hCG measurable at levels as low as 0.3 mIU per assay tube. Using 50 μl of serum sample, 6–600 mIU/ml of hCG in serum could be determined specifically with the same degree of precision as in radioimmunoassay but without sample interference with the assay. The coefficients of variation within-run and between-run were 8.6–8.9%, and 4.9– 10.7%, respectively. Values obtained with the enzyme immunoassay correlated well with those of radioimmunoassay ([unk] = 0.98, slope = 0.94, y-intercept = 10.2 mIU/ml for 75 serum samples). Results of the immunoassay of hCG levels in serial samples of serum from healthy women and patients with choriocarcinoma show that this method is useful in the clinical diagnosis of trophoblastic disease.

Competitive heterogeneous enzyme immunoassay for theophylline by flow-injection analysis with electrochemical detection of p-aminophenol

1990 ◽  
Vol 36 (4) ◽  
pp. 662-665 ◽  
Author(s):  
E P Gil ◽  
H T Tang ◽  
H B Halsall ◽  
W R Heineman ◽  
A S Misiego
Keyword(s):  
Alkaline Phosphatase ◽  
Detection Limit ◽  
Electrochemical Detection ◽  
Flow Injection ◽  
Flow Injection Analysis ◽  
General Procedure ◽  
Amperometric Detection ◽  
Enzyme Label ◽  
Commercial Kit

Abstract A competitive enzyme-linked immunoabsorbent assay based on the flow-injection amperometric detection of p-aminophenol has been investigated with use of the materials and general procedure of a commercial kit for the determination of theophylline in human serum. The antibody is immobilized on glass beads, and the enzyme label is alkaline phosphatase (EC 3.1.3.1). The high currents generated during the electrochemical detection allowed a rapid (35 min) and simple determination of theophylline throughout its therapeutic range (10-20 mg/L) and also in the subtherapeutic range (detection limit of about 80 micrograms/L).

Fully automated flow-injection system for quantifying 1,5-anhydro-D-glucitol in serum

1994 ◽  
Vol 40 (11) ◽  
pp. 2006-2012 ◽  
Author(s):  
T Tanabe ◽  
Y Umegae ◽  
Y Koyashiki ◽  
Y Kato ◽  
K Fukahori ◽  
...  
Keyword(s):  
Flow Injection ◽  
Colorimetric Detection ◽  
Enzymatic Reaction ◽  
Injection System ◽  
Gas Chromatography Mass Spectrometry ◽  
Present System ◽  
Serum Samples ◽  
Patients With Diabetes ◽  
Flow Injection System ◽  
Set Up

Abstract We have developed a flow-injection system with colorimetric detection to measure 1,5-anhydro-D-glucitol in serum. Serum samples are directly and serially injected into a clean-up column every 3.5 min to remove interferences before the enzymatic reaction. 1,5-Anhydro-D-glucitol, after being passed through the column, is oxidized by immobilized pyranose oxidase (EC 1.1.3.10), and the hydrogen peroxide produced reacts with the chromogen substrate in the presence of immobilized horseradish peroxidase (EC 1.11.1.7) to form Bindshedler's Green. The detection limit was 1.2 mumol/L (1.2 pmol). The correlation between results obtained with the present system (y) and gas chromatography-mass spectrometry (GC-MS) (x) in samples containing < 30 mumol/L 1,5-anhydro-D-glucitol, including many samples from patients with diabetes mellitus, was y = 0.975x-0.111 mumol/L (r = 0.997), which was superior to that obtained between the enzymatic and GC-MS methods. Our system needs only to be set up; it runs without any manual pretreatment, assays 17 samples/h, and shows imprecision (CV) of < 2%.

Implementation of a screening program for diagnosing open neural tube defects: selection, evaluation, and utilization of alpha-fetoprotein methodology.

1986 ◽  
Vol 32 (10) ◽  
pp. 1812-1817 ◽  
Author(s):  
R L Christensen ◽  
M R Rea ◽  
G Kessler ◽  
J P Crane ◽  
R Valdes
Keyword(s):  
Amniotic Fluid ◽  
Neural Tube ◽  
Neural Tube Defect ◽  
Gestational Age ◽  
Screening Program ◽  
Maternal Serum ◽  
Alpha Fetoprotein ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Commercial Kit

Abstract We evaluated and compared three different commercial kit immunoassays for alpha-fetoprotein (AFP) before we implemented our neural tube defect screening program. Each kit can be used with either serum or amniotic fluid. Analytical recovery ranges for AFP reference sera within each kit's standard curve limits (in kilo-int. units/L) were 97-108% (7.5-180) for the Kallestad kit, 77-101% (21.8-436) for Amersham, and 92-100% (0-177) for Hybritech. CVs, within each manufacturer's standard-curve limits, for combined intra-assay (amniotic fluid pools) and inter-assay (kit serum controls) averaged 3.6-7.3% (Kallestad), 2.4-9.3% (Amersham (y) kit results showed a correlation of r = 0.97, y = 1.05x + 5.5 kilo-int. units per liter of maternal serum (n = 66; range, 2.0-98.5). Gestational age did not influence these assay correlations. The Kallestad AFP assay demonstrated a maternal serum positivity rate of 2.9% at greater than or equal to 2.5 (n = 655) and 8.9% at less than 0.5 (n = 423) multiples of the median. All kits performed well analytically.

Evaluation of a solid-phase enzyme immunoassay for insulin in human serum.

1979 ◽  
Vol 25 (7) ◽  
pp. 1306-1308 ◽  
Author(s):  
K Kato ◽  
Y Umeda ◽  
F Suzuki ◽  
D Hayashi ◽  
A Kosaka
Keyword(s):  
Human Serum ◽  
Enzyme Immunoassay ◽  
Human Insulin ◽  
Silicone Rubber ◽  
Solid Phase ◽  
Serum Samples ◽  
Serum Factors ◽  
Insulin In Serum ◽  
Coefficients Of Variation ◽  
Sandwich Type

Abstract We describe a "sandwich-type" enzyme immunoassay for insulin in serum, in which antibody Fab'-beta-D-galactosidase conjugate and an antibody-immobilized silicone rubber solid-phase are used. The interference by serum factors with the solid-phase enzyme immunoassay can now be removed by using a buffer containing gelatin. Serum samples of 50 microL can be analyzed by the enzyme immunoassay, which is as sensitive as radioimmunoassay for human insulin. Our results correlate well with those for radioimmunoassay (r = 0.97, slope = 0.92, y-intercept = 4.6 milli-int. units /L for 181 samples). Between-assay and within-assay coefficients of variation are less than 15% over the useful ranges of the assay (5--160 milli-int. units/L).

scholarly journals Point-of-Care Quantification of Serum Alpha-Fetoprotein for Screening Birth Defects in Resource-Limited Settings: Proof-of-Concept Study (Preprint)

2020 ◽  
Author(s):  
Balaji Srinivasan ◽  
Julia L Finkelstein ◽  
David Erickson ◽  
Saurabh Mehta
Keyword(s):  
High Risk ◽  
Birth Defects ◽  
Point Of Care ◽  
Maternal Serum ◽  
Alpha Fetoprotein ◽  
Mobile Platform ◽  
Proof Of Concept ◽  
Serum Samples ◽  
Fetal Ultrasound ◽  
Serum Alpha Fetoprotein

BACKGROUND Maternal serum alpha-fetoprotein (MSAFP) concentration typically increases during pregnancy and is routinely measured during the second trimester as a part of screening for fetal neural tube defects and Down syndrome. However, most pregnancy screening tests are not available in the settings they are needed the most. A mobile device–enabled technology based on MSAFP for screening birth defects could enable the rapid screening and triage of high-risk pregnancies, especially where maternal serum screening and fetal ultrasound scan facilities are not easily accessible. Shifting the approach from clinic- and laboratory-dependent care to a mobile platform based on our point-of-care approach will enable translation to resource-limited settings and the global health care market. OBJECTIVE The objective of this study is to develop and perform proof-of-concept testing of a lateral flow immunoassay on a mobile platform for rapid, point-of-care quantification of serum alpha-fetoprotein (AFP) levels, from a drop of human serum, within a few minutes. METHODS The development of the immunoassay involved the selection of commercially available antibodies and optimization of their concentrations by an iterative method to achieve the required detection limits. We compared the performance of our method with that of commercially obtained human serum samples, with known AFP concentrations quantified by the Abbott ARCHITECT chemiluminescent magnetic microparticle immunoassay (CMIA). RESULTS We tested commercially obtained serum samples (N=20) with concentrations ranging from 2.2 to 446 ng/mL to compare the results of our point-of-care assay with results from the Abbott ARCHITECT CMIA. A correlation of 0.98 (<i>P</i>&lt;.001) was observed on preliminary testing and comparison with the CMIA. The detection range of our point-of-care assay covers the range of maternal serum AFP levels observed during pregnancy. CONCLUSIONS The preliminary test results from the AFP test on the mobile platform performed in this study represent a proof of concept that will pave the way for our future work focused on developing a mobile device–enabled quad-screen point-of-care testing with the potential to enable the screening of high-risk pregnancies in various settings. The AFP test on the mobile platform can be applied to enable screening for high-risk pregnancies, within a few minutes, at the point of care even in remote areas where maternal serum tests and fetal ultrasound scans are not easily accessible; assessment of whether clinical follow-up and diagnostic testing may be needed after a positive initial screening evaluation; and development of surveillance tools for birth defects.

Electrochemical homogeneous enzyme immunoassay of theophylline in hemolyzed, icteric, and lipemic samples

1993 ◽  
Vol 39 (7) ◽  
pp. 1432-1434 ◽  
Author(s):  
H Yao ◽  
S H Jenkins ◽  
A J Pesce ◽  
H B Halsall ◽  
W R Heineman
Keyword(s):  
Enzyme Immunoassay ◽  
Enzymatic Reaction ◽  
Normal Serum ◽  
Reaction Products ◽  
Scatter Plot ◽  
Spectrophotometric Methods ◽  
Low Concentrations ◽  
Coefficients Of Variation ◽  
Relationship Of ◽  
Homogeneous Enzyme Immunoassay

Abstract We demonstrate here an electrochemical homogeneous enzyme immunoassay for theophylline, which can be performed in hemolyzed, lipemic, and icteric samples. The assay used an unmodified Syva EMIT theophylline kit. One of the enzymatic reaction products, NADH, reacted with 2,6-dichloroindophenol (DCIP) to reduce DCIP to DCIPH2, which was detected electrochemically with flow-injection analysis. The inter- and intraassay coefficients of variation of this manual technique were &lt; 9% at theophylline concentrations of 14 to 34 mg/L. The CVs were 9-15% at low concentrations (6.3 mg/L), which is below the therapeutic range. Analytical recoveries were 91-97% for normal serum and 92-111% for hemolyzed, icteric, or lipemic sera. The measured concentrations (y) were compared with those obtained by the fluorescence polarization immunoassay (x); a scatter plot of the results showed a linear relationship of y = 1.00 x - 0.57 mg/L (r = 0.966, Sy/x = 1.51). This alternative way to measure the serum concentration of theophylline overcomes the shortcomings of spectrophotometric methods, by which it is difficult to measure theophylline in severely hemolyzed, icteric, or lipemic sera.

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