Acrosome integrity
Introduction
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Acrosome a cap like structure on the head of the spermatozoa covers 60% of the anterior portion of the sperm head.
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The morphology of the acrosome should be maintained for the sperm to undergo capacitation and acrosome reaction in the female reproductive tract for attaining fertilizing ability.
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Appreciable modifications to the structure of the plasma membrane and the outer acrosomal membrane follow after capacitation has run its course, in the form of acrosome reaction.
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Acrosome reaction consists of fusion at multiple points between the two membranes and formation of vesicles made up of fragments of two membranes.
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The sperm must be able to undergo these changes in the female reproductive tract to attain fertilizing ability by release of specific enzymes.
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The subcellular enzymes present in the acrosome facilitates disolution and penetration of the zona by the spermatozoa which will leads to the union of male and female nucleus.
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For this to happen the acrosome should be intact. Maintenance of optimum fertility depends on the acrosome being structurally and biochemically intact.
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Acrosome can be detached from sperm under the influence of different physical and chemical factors.
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Freezing and thawing can also bring about damage to the acrosome.
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Hence the acrosomal cap has received considerable attention in sperm morphology due to its importance during fertilization.
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Any damage or loss of the acrosome leads to infertility or sterility problem. Hence the evaluation of the acrosomal status gets importance.
The acrosome is evaluated by the Giemsa stain.
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Materials required
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Stock Giemsa stain
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Semen (fresh/frozen)
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Glass slides
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5% formaldehyde
Preparation of stock Giemsa Stain
Chemical
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Quantity
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Giemsa powder
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1 gm
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Glycerol
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60 ml
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Methanol
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66 ml
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Weigh the Giemsa powder, put in a pestle and mortar.
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The glycerol is added slowly and grind it well. Then add methanol slowly and grind it.
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Finally the solution is filtered and stored in an amber colored bottle.
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Preparation of working Giemsa solution Preparation of Sorenson’s Buffer Preparation of 5% formaldehyde
Chemical
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Quantity
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Na2HPo4.2H2O (21.682 gm/500 ml)
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200 ml
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KHPo4H2O (22.254 gm/500 ml)
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80 ml
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Stock buffer
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280 ml
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Staining procedure and result interpretaion
Procedure (Video demonstration: Giemsa staing procedure Part 1 & Giemsa staing procedure Part 2)
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A drop of diluted semen (1:5 to 1:10 in 2.9 per cent sodium citrate) is kept on a clean, grease free, pre-warmed glass slide and is dried in air. The frozen thawed semen needs no dilution.
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The slide is immersed in 5% formaldehyde f or 30 minutes for fixing semen smear.
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The slide is washed in running tap water and dries it.
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The slide is immersed in working Giemsa solution for 3 hours at 37 ⁰ C.
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Finally wash the slide in running tap water and dry it.
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Examine the slide under oil immersion of phase contrast microscope and count 200 sperms.
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The acrosome based on the morphology is classified as follows
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S.No.
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Defect
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Picture
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1
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Normal acrosome
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2
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Partially lost acrosome
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3
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Completely lost acrosome
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4
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Ruffled/wrinkled acrosome
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5
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Knobbed acrosome
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6
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Diadem defect
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7
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Detached galea capitis
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8
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Apical notch/ridge
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Various acrosomal defects are given below.
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Last modified: Monday, 4 June 2012, 10:05 AM