When infertility occurs, both spouses are tested to find the cause. One of the essential components is laboratory testing. For men, besides the sperm vitality, concentration and abnormalities that you know about, what other tests can be done on the semen that you don’t know about?

I. Routine Semen Analysis

These include semen volume, appearance, liquefaction time, acidity, sperm count, motility, viability and morphology.

(1) Volume: a normal male ejaculate volume of ≧ 1.5ml, usually 1.5 to 6ml, of course under the premise of abstinence for 3 to 7 days. Too little semen volume is not conducive to sperm traveling through the vagina into the uterus and fallopian tubes, which affects conception. Too much semen volume and the sperm is diluted, resulting in low sperm concentration, which is also not conducive to fertility.

(2) Appearance: Normal semen is a milky white, off-white, homogeneous, semi-fluid liquid. If the semen is pale yellow, it is more likely that the abstinence period is too long; if it is red or coffee-colored, there may be inflammation of the prostate or seminal vesicles.

(3) Liquefaction time: Liquefaction time refers to the time it takes for normal semen to change from jelly-like to liquid after ejaculation. The liquefaction time of normal human semen is usually 15-30 minutes. If it takes more than 60 minutes and is still jelly-like, it is a pathological condition and is called semen non-liquefaction. The cause of non-liquefaction is not completely clear. It may be due to insufficient fibrinolytic enzymes secreted by the prostate gland due to seminal vesiculitis or prostatitis, or the deficiency of some trace elements such as magnesium and zinc, etc.

(4) Semen odor: Semen has a distinctive fishy odor due to the oxidation of sperm ammonia.

(5) pH: The normal value is 7.2-8.0. pH>8.0 is common in acute epididymitis, seminal vesicle adenitis, prostatitis, etc.; pH<7.2 is common in congenital vas deferens obstruction, congenital seminal vesicle or epididymis dysplasia; pH≤6.0 is associated with seminal vesicle and epididymis dysplasia.

(6) Sperm concentration and total sperm count: Sperm concentration is generally ≥15×106/ml; less than this value is oligospermia. The total sperm count is usually ≥39×106/1 ejaculation. If the sperm concentration is too low or the total number of sperm is too low, it often affects the female partner’s ability to conceive.

(7) Sperm vitality: Sperm vitality refers to the motility of sperm. The WHO classifies it into three levels to determine. pr indicates fast linear forward motion of sperm, straight line motion; np indicates no forward motion of sperm, in situ motion; im indicates inability of sperm to move, no motion. If normal, PR spermatozoa ≥ 32%, if PR spermatozoa < 32%, it is weak spermatozoa.

(8) Sperm survival rate: Sperm survival rate refers to the proportion of surviving sperm in the total number of sperm. The sperm survival rate of normal people should be ≧ 58%. Low sperm survival rate is common in prostatitis, vesiculitis, varicocele, etc.

(9) sperm morphology: the proportion of normal human sperm in normal form should be ≥ 4%; if the proportion of deformed sperm > 96%, may cause fertility disorders, and even lead to the female embryo stop developing or spontaneous abortion, etc.. Generally, sperm Pap staining must be performed to accurately calculate the percentage of sperm malformation.

2.Semen biochemical examination

Eminal plasma biochemical tests: including tests for neutral A glucosidase, elastase, seminal plasma zinc, fructose, acid phosphatase, citric acid and sperm lactate dehydrogenase isoenzyme.

(1) Neutral-a glucosidase assay: mainly secreted by epithelial cells of epididymis, it is a specific enzyme and marker enzyme of epididymis, which can catalyze the decomposition of sugar component of protein or oligosaccharide to provide suitable energy for sperm maturation, and this enzyme can be used as an evaluation index of secretory function of epididymis. Meanwhile, combined with seminal plasma fructose test, it can localize the obstruction site in patients with obstructive azoospermia, which greatly improves the diagnosis rate of obstructive azoospermia.

(2) Seminal plasma elastase assay: Seminal plasma elastase is one of the means to detect inflammation, and can be used as an indicator for the diagnosis and healing of dormant reproductive tract infections.

(3) Seminal plasma zinc assay: Zinc in semen comes mainly from the prostate gland, and its content is more than 100 times higher than that of blood zinc. It is now believed that seminal plasma zinc is an important factor in maintaining sperm viability. Zinc has a reversible inhibitory effect on sperm acrosome enzymes. When sperm enter the cervical mucus, zinc-binding proteins in the mucus can lower the zinc level, leading to the activation of acrosome enzymes, allowing sperm to pass through the zona pellucida and unite with the egg. In addition, seminal zinc is one of the indicators of prostate gland function.

(4) Seminal plasma fructose determination: Fructose in seminal plasma is produced by enzymatic conversion of blood glucose and is secreted by the seminal vesicle gland. It is the energy source for sperm activity. Since the seminal vesicle gland is sensitive to androgen stimulation and the secretion of fructose is controlled by androgens, seminal plasma fructose measurement can be used to monitor the function of seminal vesicles and testicular interstitial cells, and also helps to diagnose the cause of azoospermia.

(5) Seminal plasma acid phosphatase assay: Acid phosphatase, which catalyzes the hydrolysis of phosphate bonds, is present in all tissues of the body and is particularly abundant in the prostate. The activity of this enzyme may be significantly altered in prostate disease.

(6) Seminal plasma citric acid determination: Seminal plasma has a high citric acid content, almost all of which comes from the prostate. Citric acid is involved in maintaining the osmotic pressure of seminal plasma and is associated with sperm vitality. The measurement of citric acid content in seminal plasma is also of some reference value for the detection of prostate function and male sexual function.

(7) Lactate dehydrogenase isoenzyme X (LDH-X) is located in mature testes and spermatozoa of mammals and birds and is a sperm-specific LDH whose synthesis is regulated in relation to androgens, involved in sperm energy metabolism, important for spermatogenesis and maturation, and can be used as a specific indicator of male fertility.

3.Semen pathogenic microbiological examination

Microbial infections of the male reproductive system can damage the reproductive organs and affect spermatogenesis, maturation and transport, resulting in decreased semen quality and male infertility. These pathogenic microorganisms include mycoplasma, chlamydia, gonococcus, bacteria, etc., which can be tested by semen culture, gene amplification and other methods.

4.Semen Anti-Sperm Antibody Test

Anti-sperm antibodies produced by the male partner due to reproductive tract infection, injury, surgery, etc., can have a significant impact on sperm production and sperm movement, which may affect fertilization, fertilized egg implantation and development, and affect fertility.

5.Activity of Sperm Acrosome Enzymes and Assay of Induced Sperm Acrosome Reaction

Sperm acrosome enzymes are present in the inner membrane of the sperm acrosome and in the equatorial part of the membrane, usually in an inactive form, and only when the sperm head enters the zona pellucida of the egg is the acrosome zymogen activated as an acrosome enzyme. The acrosome reaction refers to a series of changes that occur when the sperm are energized and meet the egg. It usually occurs after the sperm has joined the zona pellucida of the egg and before the sperm penetrates the egg membrane and fertilizes the egg, and is one of the necessary parts of natural fertilization of the egg. Therefore, even if the spermatozoa are highly energetic and numerous, if the acrosome enzyme activity of the sperm is insufficient or the acrosome reaction is insufficient, it still cannot naturally impregnate the woman.

6.Sperm DNA damage detection

Sperm DNA integrity test is a new indicator to respond to male fertility and one of the common important reasons affecting male fertility, which is influenced by multiple factors such as genetics, environment and lifestyle habits, and is closely related to male infertility and recurrent miscarriage of spouse. It has been shown that sperm DNA breakage index (DFI) is higher in infertile men with abnormal semen parameters (oligospermia, weak sperm, etc.) than in men with normal fertility, and DFI is also increased in idiopathic infertility patients with normal semen parameters; when DFI ≥ 30%, the chance of natural pregnancy is lower; in assisted reproduction techniques, sperm DNA damage is also one of the important factors affecting its outcome. Therefore, DFI is clinically important for the assessment of male fertility and the selection of assisted reproductive technologies.
  DFI (DNA fragmentation index)
  ≤15%: good sperm nuclear DNA integrity;
  >15%, <25%: moderate sperm nuclear DNA integrity;
  ≥25%: poor sperm nuclear DNA integrity.
  HDS (HighDNAStainability): represents immature sperm in semen, normal value <15%, usually combined with DFI assessment to increase the accuracy of fertility assessment.

7.Seminal Plasma 8-Hydroxydeoxyguanosine Assay

8-hydroxy-2-deoxyguanosine (8-OHdG) is a DNA damage marker that can be used to determine the extent of oxidative damage to sperm DNA by reactive oxygen species and the cause of abnormal sperm formation.
Studies have shown that when 8-OHdG is elevated in sperm DNA, the head malformation rate increases significantly. The 8-OHdG in sperm DNA was negatively correlated with the normal sperm rate. The results suggested that there was a correlation between 8-OHdG in sperm DNA and semen quality, indicating that the decrease in semen quality was related to the damage of sperm DNA, and endogenous and exogenous reactive oxygen species were one of the possible causes.

8.Seminal plasma free miRNA detection

Seminal plasma-free RNA is an extracellular RNA found in seminal plasma and originates from both testes, epididymis, seminal vesicles and prostate. It is clinically important as a non-invasive method for classifying azoospermia and predicting spermatogenesis.

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