Sperm Function Test
Nowadays male infertility has increased and counts near about 40% of infertility among couples. A basic semen analysis is the main first step to evaluate the male infertility. Under the routine Semen analysis procedure, semen is evaluated for its volume viscosity and appearance. semen is analysed as per the WHO guidelines to determine its concentration and motility. Vitality can be determined by the Eosin Nigrosin stain and the morphology by preparing the smear and staining with papanicolaou stain. These all above-mentioned measures are the assessment of its structure. The chromatin structure of the sperm is highly compact and stable. The abnormal development of the DNA loop can cause sperm DNA defects. The chromatin integrity of the sperm is directly correlated with the fertility of men and it has been found that a normal Semen analysis can also have a higher DFI. The semen with less amount of DNA fragmentation is associated with less time to achieve natural pregnancy. Therefore several tests are now used to assess the functional ability of the sperm.
1. Acidic aniline blue stain
The spam with immature nuclear chromatin is stained blue while those with mature nuclei do not take up the stain.
2. Chromomycin A3 assay
This method reveals poly pack chromatics.
3. Acridine Orange essay
The spermatozoa emitting green colour indicates Intact DNA where the fragmented DNA looks yellow, orange or red colour.
4. The comet assay
It is a second generation sperm DNA fragmentation analysis test that is very sensitive to estimate both single and double stranded breaks; it can quantify the actual amount of DNA damage per sperm.
5. TUNEL assay
This is an extremely sensitive and reliable essay to be stored for a longer time after fixing but it is a time consuming method.
6. The sperm chromatin essay
This is an accurate estimation of the percentage of DNA damage raisins are analysed with the help of a software displaying the ratio of green and red sperm. The spam with the fragmented DNA gives the red emission and sperm with the intact structure gives the green emission.
7. sperm chromatin dispersion assay
It is based on interpreting the difference between the fragmented and the non fragmented DNA in the spermatozoa depending on the formation of Halos. It is a kit based method. The spermatozoa with the intact DNA shows big halos while the spermatozoa with the fragmented DNA shows minimal or no Halos. The Halos are visualised either by fluorescent stain or by bright field microscopy stain.