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LH
Luteinizing hormone (LH) is a glycoprotein consisting of two subunits with a molecular mass of 30,000 daltons. The similar to other pituitary hormones [follicle stimulating hormone (FSH), thyroid stimulating hormone (TSH) and chorionic gonadotropin (HCG)] while the The 15% and 30%. The clinical usefulness of the measurement of luteinizing hormone (LH) in ascertaining the homeostasis of fertility regulation via the hypothalamic -pituitary - gonadal axis has been well established (1,2). In addition, the advent of in vitro fertilization (IVF) technology to overcome infertilityassociated problems has provided the impetus for rapid improvement in LH assay methodology from the technically demanding bioassay to the procedurally simple and rapid immunoenzymometric assays.
LH ELISA:
Immunoenzymatic colorimetric method (ELISA) for quantitative determination of LH in serum.
Principle of the Assay:
In this method, LH standards, patient specimens and/or controls (containing the native antigen) are first added to streptavidin coated wells. Biotinylated monoclonal and enzyme labeled antibodies are added and the reactants mixed: these antibodies have high affinity and specificity and are directed against distinct and different epitopes of LH. Reaction between the various LH antibodies and native LH occurs in the microwells without competition or steric hindrance forming a soluble sandwich complex. The interaction is illustrated by the following equation:
ka |
BtnAb(m) = Biotinylated Monoclonal Antibody (Excess Quantity)
AgLH = Native Antigen (Variable Quantity)
EnzAb(p) = Enzyme labeled Antibody (Excess Quantity)
EnzAb(p)-AgLH-BtnAb(m) = Antigen-Antibodies Sandwich Complex
ka = Rate Constant of Association
k-a = Rate Constant of Dissociation
Simultaneously, the complex is deposited to the well through the high affinity reaction of streptavidin and biotinylated antibody. This interaction illustrated below:
EnzAb -AgLH-BtnAb(m) + StreptavidinC.W. -> Immobilized complex |
Streptavidin C.W. = Streptavidin immobolized on well.
Immobilized complex = Antibodies-Antigen sandwich bound.
After equilibrium is attained, the antibody-bound fraction is separated from unbound antigen by decantation or aspiration. The enzyme activity in the antibody-bound fraction is directly proportional to the native antigen concentration. The activity of the enzyme present on the surface of the well quantitated by reaction with a suitable substrate to produce colour. By utilizing several different calibrators of known antigen values, a dose response curve can be generated from which the antigen concentration of an unknown can be ascertained.
Specific performance characteristics:
Intra Assay Variation:
Within-run precision was determined by replicate determinations of three different control sera in one assay. The within-assay variability is shown below:
Serum Sample | 1 | 2 | 3 |
Number of Replicates | 20 | 20 | 20 |
Mean LH (mIU/ml) | 1.0 | 8.1 | 18.9 |
Standard Deviation | 0.08 | 0.064 | 1.15 |
Coefficient of Variation (%) | 8.0 | 7.9 | 6.1 |
Inter Assay Variation:
Between-run precision was determined by replicate measurements of three different control sera in several different assays. The between-assay variability is shown below:
Serum Sample | 1 | 2 | 3 |
Number of Replicates | 10 | 20 | 20 |
Mean LH (mIU/ml) | 1.1 | 8.3 | 18.9 |
Standard Deviation | 0.013 | 0.75 | 1.60 |
Coefficient of Variation (%) | 11.8 | 9.0 | 8.5 |
Analytical Sensitivity:
The minimal detectable concentration of LH by this assay is estimated to be 0.5 mIU/ml.
Accuracy:
Correlation between the NovaTec LH kit and the well-established RIA method (which used standards calibrated against the previous 1st IRP 68/40) gave the following results:
r = 0.940
n = 157
Order information:
ELISA | Number of Determinations | Product Number |
LH | 96 | DNOV030 |
