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Testis Biopsy


Testis biopsy was first reported by Hotchkiss and Engle at The New York Hospital-Cornell Medical Center in the late 1930s. The primary purpose of testis biopsy is to distinguish between obstructive azoospermia and primary seminiferous tubular failure. It is indicated in men who are azoospermic with testis of normal size (greater than 15cc in volume), normal consistency, palpable vasa deferentia and normal serum FSH levels. Azoospermia in men with small soft testes and FSH levels elevated more than twice normal is always due to primary germ cell failure and biopsy may be helpful to rule out intratubular germ cell neoplasm (carcinoma-in-situ). However, biopsy of men with presumed non-obstructed azoospermia should only be performed if cryopreservation of the tissue is possible at the same time. Men with borderline FSH levels and slightly soft small testes probably have testicular failure and biopsy should only be performed in these men when absolute confirmation of the diagnosis is required.

We no longer perform biopsy on men without palpable vasa. We have biopsied a dozen such men and all of these biopsies have revealed virtually normal spermatogenesis. The presence or absence of spermatozoa is the ultimate and only determinant of whether the patient is a candidate for ICSI (Intracytoplasmic Sperm Injection) using sperm retrieval. Also, testis biopsy should almost always be performed bilaterally. Small firm testes often have surprising good spermatogenesis. Biopsies of even large healthy testis sometimes reveal maturation arrest.

Technique of Open Testis Biopsy

Once the decision has been made to perform testis biopsy, it is the obligation of the surgeon to provide a tissue sample adequate for good pathological evaluation using a technique that minimizes trauma to the specimen and prevents injury to the epididymis and testicular blood supply. It also need to coordinate the time schedule with the IVF embrology lab if performing of TESA at the same time.

With the assistant stretching the scrotal skin tightly,over the anterior surface of the testis and assuring that the epididymis is posterior, bilateral 5 mm transverse scrotal incisions provide good exposure with a minimum of scrotal skin bleeding. The incision is carried through the skin and dartos muscle and the tunica vaginalis is opened. Bleeders on the edges of the tunica vaginalis are cauterized. The tunica albuginea is exposed and transfixed with a 4-0 catgut suture armed with a small tapered needle (cutting needles often cause more bleeding than the suture prevents). All bleeders are coagulated prior to incising the tunics to prevent saturation of the biopsy with blood. A 3 to 4 mm incision is made with the point of the scalpel into the tunic and a pea-sized sample of seminiferous tubules excised with a razor sharp iris scissors and deposited directly into either Bouins, Zenkers, or collidine buffered glutaraldehyde solution. Formalin fixation results in severe distortion of the testicular histology, making these specimens almost impossible to read. It should never be used for testis biopsy. A second specimen is placed on a slide, a drop of saline or Ringer's is added, and the specimen is squashed under a coverslip to allow sperm to leave the tubules. The squash prep is examined under a microscope using phase contrast. The presence of sperm with tails, especially motile, on the squash prep is almost 100% predictive of obstruction. The tunics are closed with a running 5-0 polypropylene monofflainent suture. Skin sutures are not required. The wounds need only be covered with Bacitracin ointment and a fluff type dressing held in place with snug scrotal supporter. Antibiotics are unnecessary. Tylenol or Tylenol with codeine provides adequate analgesia.

Technique of Percutaneous Testis Biopsy

Percutaneous testis biopsy using a Tru-Cut type of devise has been performed as an office procedure under local anesthesia. It has been used fo evaluation of both histology and cytology. This blind biopsy procedure could result in unintentional injury to either the epididymis or testicular artery coursing under the surface of the tunica albuginea. In addition, we have often found that specimens obtained in this way often contain only three to six tubules with poorly preserved architecture. Specimens obtained in this way can be used to extract sperm for ICSI in case of obstruction.

Technique of Percutaneous Testicular Fine Needle Aspiration (TFNA)

Aspiration with a fine gauge needle may be less risky and painful than percutaneous biopsy. Evaluation of such material with flow cytometry offers promise as a useful diagnostic tool. Only a few studies of such techniques have been reported. Until standards for the evaluation of aspirated material are well established, open testis biopsy is the diagnostic procedure of choice. Fresh unfixed testis biopsy materials should be examined in the operating room to determine sperm are presented and whether they are motile. Sperm can be obtained this way for ICSI in case of obstruction.


Examination of properly fixed biopsy material by light microscopy provides most of the information necessary to make an accurate diagnosis. Mature spermatids and sperms are easy to differentiate from other germ cell elements. An average of at least twenty mature spermatids and/or sperms per/tubule from at least 10 counted tubules predicts a total sperm count of at least 10 million. Azoospermic or severely oligospermic men with spermatid counts considerably higher than this are almost certain to be obstructed. Electron microscopic examination of testicular biopsy material yields little additional information on the cause of most testicular disorders. The electron microscope is useful in the diagnosis of rare disorders such as Kartageners Syndrome, in which the dynein side arms are absent from sperm tails. Such diagnoses can be more easily made by a thorough history and physical examination. Abnormalities of sperm head shape, such as round heads or absent acrosomes are clearly defined with the electron microscope, but can also be diagnosed with routine staining and light microscopy. Flow cytometric evaluation of testis biopsies can detect different patterns of nucleic acid content and distribution. Maturation beyond the meiotic stage can readily be detected with flow cytometry. This technique might prove to be a useful screening tools when standards have been validated.


When performed with care, testis biopsy is associated with few complications. The most serious misadventure associated with testis biopsy would be biopsy of the wrong structure, in particular biopsy of the epididymis. If examination of the biopsy material reveals epididymis with sperm within the epididymal tubule, obstruction of the epididymis at the site of the biopsy is certain. If, however, biopsy of the epididymis reveals the absence of sperm in the tubules, no serious harm has been done, and the patient is either obstructed above the level of the biopsy or has primary seminiferous tubular failure and is not making sperm. Large hematomas can result from testis biopsy if the testicular artery on the surface of the tunica albuginea has been injured. If this injury is not recognized, hematomas can grow to frightening size and require drainage. Leaving the tunica vaginalis and tiny skin wounds open will almost always prevent hematoma formation, at the very worst, the patient would have bloody bandages and the bleeding would eventually subside. Because of the rich testicular blood supply, wound infection is extremely rare in the absence of hematoma. Antibiotics are not routinely indicated after testis biopsy.Testicular atrophy might result if the testicular artery is injured during the course of a blind percutaneous biopsy or a biopsy under local anesthesia using a blind cord block.Placement of the biopsy material in the incorrect fixative such as formalin, or inadequate sampling of the testicular tissue, renders the biopsy use less and it would then have to be repeated.

Center for Male Reproductive Medicine & Microsurgery Weill Cornell Medicine
525 E 68th Street
New York, NY 10065