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. 2022 Jun;57(6):678-684.
doi: 10.1111/rda.14100. Epub 2022 Mar 4.

Comparison of two staining techniques on the manual and automated canine sperm morphology analysis

Paulina Surmacz  1 , Anna Niwinska  1 , Ewa Kautz  2 , Slawomir Gizinski  1 , Ricardo Faundez  3
Affiliations

Comparison of two staining techniques on the manual and automated canine sperm morphology analysis

Paulina Surmacz et al. Reprod Domest Anim. 2022 Jun.

Abstract

Detailed and direct analysis of semen, including sperm morphology, enables a diagnosis of male fertility. This study aimed to describe an economical and verified protocol for canine spermiograms and compare the effectiveness of Sperm Stain® and Sperm Blue® (Microptic, Spain) in veterinary practice. Sperm assessment was conducted manually, using a standard optical microscope, and via computerized semen analysis using the SCA® CASA (Sperm Class Analyzer® CASA System-MICROPTIC, Spain). This study showed that Sperm Blue® is a better solution for computerized sperm quality analysis of healthy dogs. At the same time, Sperm Stain® turned out to be more helpful in identifying specific morphological defects of sperm. Automated canine sperm morphology analysis worked better with Sperm Blue stain, but Sperm Stain simplified manual evaluation of various organelles' defects. Standard, manual examination is more error-prone for an inexperienced andrology technician, but it seems to be still a gold standard technique for canine sperm assessment.

Keywords: CASA; Sperm Blue®; Sperm Stain®; andrology; dog spermiogram; sperm morphology.

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Conflict of interest statement

The authors whose names are listed immediately below certify that they have NO affiliations with or involvement in any organization or entity with any financial interest (such as honoraria; educational grants; participation in speakers' bureaus; membership, employment, consultancies, stock ownership or other equity interest; and expert testimony or patent‐licensing arrangements) or non‐financial interest (such as personal or professional relationships, affiliations, knowledge or beliefs) in the subject matter or materials discussed in this manuscript.

Figures

FIGURE 1
FIGURE 1
Defects in the heads, acrosomes, insertions and tails of canine spermatozoa. Authors’ rendition. Sperm Blue® (1,2,3,6,7,8,9,10,12,13,14,15,16,18,19,20,23,24,25,26,27,28,31,33,36,37) Sperm Stain® (4,5,11,17,21,22,29,30,32,34,35). 1 – ruptured acrosome; 2 – detached head; 3 – midpiece defect; 4 – acrosome and midpiece atrophy; 5 – acrosome stratification; 6 – micro‐sperm head; 7 – small sperm head; 8 – amorphous head, abnormal midpiece angle; 9 – amorphous head; 10 – wrapping of midpiece; 11 – an absence of acrosome; 12 – pear‐shaped head; 13 – granules in sperm head;14 – pear‐shaped head and absence of well‐defined acrosome; 15 – micro‐round head, 16 – head at the midpiece erosion; 17 – double‐headed sperm; 18 – cyst at the apex of the sperm acrosome; 19 – granules in the sperm head; 20 – double‐headed, amorphous sperm; 21 – tapered head; 22 – double‐headed sperm; 23 – vacuoles in the sperm head; 24 – cyst at the apex of the head; 25 – paintbrush head; 26 and 35 – rolling of sperm tail; 27 – two tails; 28 and 36 – acrosome swelling and rolling of tail; 29 and 34 – Dag defect; 30–2 – double sperm; 31 and 32 – the presence of distal cytoplasmic droplet; sperm with short tail; 33 – short tail; 37 – normal sperm head
FIGURE 2
FIGURE 2
Diagram of a smear preparation. Authors’ rendition
FIGURE 3
FIGURE 3
Comparison of subjective, manual evaluation of Sperm Stain® and Sperm Blue® stained samples. Normal/abnormal spermatozoa
FIGURE 4
FIGURE 4
Summary of subjective morphological evaluation of spermatozoa. Comparison of Sperm Stain® and Sperm Blue® for different teratozoospermic characteristics
FIGURE 5
FIGURE 5
Comparison of automatic Sperm Stain® and Sperm Blue® staining analysis in CASA 5.4.0.0 system. Normal/abnormal spermatozoa
FIGURE 6
FIGURE 6
Comparison of automatically detected morphological defects results from the same samples stained with Sperm Stain® and Sperm Blue®. Analysis in CASA 5.4.0.0 system
FIGURE 7
FIGURE 7
Summary of sperm morphological defects. Comparison of Sperm Stain® and Sperm Blue® staining. Analysis in CASA system
FIGURE 8
FIGURE 8
Analysis of CASA 5.4.0.0 SCA Research Edition – Morphology module (Microptic, Spain) A‐F Sperm Stain sperm staining; G‐M Sperm Blue staining. Authors’ rendition
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