Enzyme Immunoassay for the Quantitative Determination of Luteinizing Hormone (LH) Concentration in Human Serum

for in vitro diagnostic use

Product Description
Luteinizing hormone (LH) is produced in both men and women from the anterior pituitary gland in response to luteinizing hormone-releasing hormone (LH-RH or Gn-RH), which is released by the hypothalamus. LH, also called interstitial cell-stimulating hormone (ICSH) in men, is glycoprotein with a molecular weight of approximately 30,000 daltons. It is composed of two noncovalently associated dissimilar amino acid chains, alpha and beta. The alpha chain is similar to that found in human thyroid-stimulating hormone (TSH), follicle-stimulating hormone (FSH), and human chorionic gonadotropin (hCG).

The differences between these hormones lie in the amino acid composition of their beta subunits, which account for their immunological differentiation.

The basal secretion of LH in men is episodic and has the primary function of stimulating the interstitial cells (Leydig cells) to produce testosterone. The variation in LH concentrations in women is subject to the complex ovulatory cycle of healthy menstruating women, and depends on a sequence of hormonal events along the gonado-hypothalamicpituitary axis. The decrease in progesterone and estradiol levels from the preceding ovulation initiates each menstrual cycle. As a result of the decrease in hormone levels, the hypothalamus increases the secretion of gonadotropin-releasing factors(GnRF), which in turn stimulates the pituitary to increase FSH production and secretion. The rising FSH levels stimulate several follicles during the follicular phase, one of these will mature to contain the egg. As the follicle develops, estradiol is secreted, slowly at first, but by day 12 or 13 of a normal cycle increasing rapidly. LH is released as a result of this rapid estradiol rise because of direct stimulation of the pituitary and increasing GnRF and FSH levels. These events constitute the pro-ovulatory phase. Ovulation occurs approximately 12 to 18 hours after the LH reaches a maximum level. After the egg is released, the corpus luteum is formed which secretes progesterone and estrogen feedback regulators of LH. The luteal phase rapidly follows this ovulatary phase, and is characterized by high progesterone levels, a second estradiol increase, and low LH and FSH levels. Low LH and FSH levels are the result of negative feedback effects of estradiol and progesterone on the hypothalamic-pituitary axis. After conception, the developing embryo produces hCG, which causes the corpus luteum to continue producing progesterone and estradiol. The corpus luteum regresses if pregnancy does not occur, and the corresponding drop in progesterone and estradiol levels results in menstruation. The hypothalamus initiates the menstrual cycle again as a result of these low hormone levels.

Patients suffering from hypogonadism show increased concentrations of serum LH. A decrease in steroid hormone production in females is a result of immature ovaries, primary ovarian failure, polycystic ovary disease, or menopause; in these cases, LH secretion is not regulated. A similar loss of regulatory hormones occurs in males when the tests develop abnormally or anorchia exists. High concentrations of LH may also be found in primary testicular failure and Klinefelter syndrome, although LH levels will not necessarily be elevated if the secretion of androgens continues. Increased concentrations of LH are also present during renal failure, cirrhosis, hyperthyroidism, and severe starvation. A lack of secretion by the anterior pituitary may cause lower LH levels. As may be expected, low levels may result in infertility in both males and females. Low levels of LH may also be due to the decreased secretion of GnRH by the hypothalamus, although the same effect may be seen by a failure of the anterior pituitary to respond to GnRH stimulation. Low LH values may therefore indicate some dysfunction of the pituitary or hypothalamus, but the actual source of the problem must be confirmed by other tests.

In the differential diagnosis of hypothalamic, pituitary, or gonadal dysfunction, assays of LH concentration are routinely performed in conjugation with FSH assays since their roles are closely interrelated. Furthermore, the hormone levels are used to determine menopause, pinpoint ovulation, and monitor endocrine therapy.

Principle
The LH Quantitative Test is based on a solid phase enzymelinked immunosorbent assay (ELISA). The assay system utilizes a mouse monoclonal anti-á-LH antibody for solid phase (microtiter wells) immobilization and a mouse monoclonal anti-â-LH antibody in the antibody-enzyme (horseradish peroxidase) conjugate solution. The test sample is allowed to react simultaneously with the antibodies, resulting in LH molecules being sandwiched between the solid phase and enzyme-linked antibodies.
After a 45-minute incubation at room temperature, the wells are washed with water to remove unbound-labeled antibodies. A solution of TMB Reagent is added and incubated for 20 minutes, resulting in the development of a blue color. The color development is stopped with the addition of Stop Solution, and the color is changed to yellow and measured spectrophotometrically at 450nm.
The concentration of LH is directly proportional to the color intensity of the test sample.

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