Neuropathology In the Department of Pathology

Procedures For Nerve and Muscle Biopsies

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Nerve Biopsy   >Surgeon | >Pathologist
Muscle Biopsy >Surgeon | >Pathologist


Information for Surgeon

Sural nerve biopsy at ankle level:

Compression of calf causes the lesser saphenous vein, which is a very reliable landmark for locating the sural nerve, to distend and to become visible or palpable in the trough between the external malleolus and Achilles tendon.  The sural nerve is usually located medial to the vein.

Infiltrate skin and subcutaneous tissue with 10 cc Lidocaine (0.5%) behind the external malleolus.

Place incision along course of short saphenous vein.

Divide Scarpa’s fascia and retract vein, exposing nerve usually located medial to the vein. 

Secure bleeders and infiltrate the most proximal end of the exposed nerve with 0.2 cc Lidocaine.

Free nerve from surrounding soft tissue, preferably with sharp dissection. (Avoid rubbing nerve with gauze – i.e., blunt dissection – and vigorous stretching, to prevent artifacts, especially of the myelin sheath.)

Ligate nerve at site of Lidocaine injection and cut nerve distal to ligature.

2.5 cm length of nerve is ample for both light and electron microscopy.

Wrap biopsy in gauze slightly moistened with NORMAL SALINE. The gauze should not be damp-soaked wet.  Do not drop specimen in saline. Place specimen in Petri dish and cover. Call Neuropathology personnel immediately to pick up specimen.

If surgery is done in the main operating rooms, call Neuropathology at 3-6041 and take specimen to ADULT HOLDING and Neuropathology personnel will pick it up.

If surgery is done in the ASU Treatment Room, we will pick up specimen outside the room.

Information for Pathologists

Regional nerve biopsies:

  1. Please telephone us (650-723-6041) at least one day before the biopsy so that we can prepare to appropriately process the specimen.

  2. If the nerve is delivered fresh from a local institution, use saline moistened gauze, place in a water-tight container and place on wet ice prior to expedited transport to Stanford Neuropathology.

Biopsies from outside the SUMC or regional area:

  1. Isotonic glutaraldehyde is highly desirable and may be prepared according to the following:
    10 cc of 0.1 molar cacodylate buffer
    +    10 cc 8% glutaraldehyde
    +    10 cc distilled water. 

    If this is not possible, use buffered 2.5% glutaraldehyde.

  2. About one third of the nerve biopsy is carefully excised from the specimen and placed in formalin for paraffin embedding.

  3. Do not freeze any of the nerve.

  4. The remainder of the specimen must be placed in the isotonic glutaraldehyde, if available, after first allowing the nerve to adhere to dry stiff filter paper (index card, blotting paper) thus kept in a straight position, ideally at 4 degrees C, while the specimen undergoes fixation. 

  5. If the biopsy originates from institutions within the greater San Francisco Bay Area region, and cannot be delivered within 30 minutes, it is highly preferable that it be divided for fixation as specified above. If glutaraldehyde is not available at your institution, please place the entire specimen in formalin, and we will post-fix a portion in glutaraldehyde.

  6. The specimens should be shipped for further processing to the Neuropathology Laboratory at:

Stanford University Medical Center
Neuropathology, Room R-241
300 Pasteur Drive
Palo Alto, CA 94305-5324



Information for Surgeon

  1. We request two days notice to schedule receipt of the specimen at the Stanford University Medical Center. Call your hospital pathologist in advance of surgery so that he may arrange transportation. If the procedure is done outside the Stanford University complex but within the greater San Francisco Bay Area region, the specimen must reach our laboratory within 6 hours after the biopsy. In order to give us sufficient time to process the specimen, the biopsy should reach us no later than 3 PM.

  2. Preferred biopsy sites:
    • Biceps, deltoid, vastus lateralis, quadriceps, or as specified by the neurologist.
    • If the patient is suspected to have an inflammatory myopathy, submit in addition a piece of fascia.
    • If a metabolic myopathy is suspected, take an additional piece for biochemical studies.
    • If the symptoms are intermittent perform the biopsy while the patient is experiencing a relapse.

  3. Surgical technique:
    • Local infiltration with lidocaine without epinephrine. Donot infiltrate the muscle.
    • Avoid muscle that has had EMG manipulation or severely wasted muscle, or muscle that is not affected or normal.
    • Avoid electrocautery completely in opening the muscle and all aspects of the biopsy until after the specimen has been removed. Cautery causes severe artifact, not only by rendering the cauterized portions uninterpretable, but often results in contraction artifact throughout the specimen.
    • Take the biopsies from the belly of the muscle, avoiding when possible subfascial and myotendonous areas.
    • Note: Isometric muscle biopsy clamps are not necessary in the removal of the biopsies, and should be specifically avoided in infant or young pediatric biopsies. Regardless of whether clamps are used, two pieces are required.
    • Please obtain two intact pieces of muscle, each approximately 1-2 cm in length X 0.5-0.8 cm. in thickness.
    • Wrap the specimens in gauze that has been lightly dampened with saline.  Never immerse a specimen in saline or fixative of any kind before submitting to the pathologist.
    • Small fragments or multiply incised biopsies are often inadequate for diagnosis.

Information for Pathologist 

  1. The best portion of the muscle should be snap frozen by the following method:
    • Upon receipt of the specimen, and using a fresh blade carefully dissect a cylindrical piece of muscle approximately 1 cm. in length and up to 0.5 cm. in diameter from the specimen.
    • The portion not frozen may be used for formalin fixation, however:
    • The frozen portion is the most important part of the tissue for diagnosis and biochemical testing, therefore formalin fixation may be omitted if the specimen is inadequate.
    • If the biopsy is of adequate size, 2 or 3 pieces should be frozen in a similar manner. 
    • The portions for freezing should be placed in a plastic mold such as is used to prepare blocks for frozen sections but without OCT or similar freezing embedding media and snap frozen by the method employed at the referring institution. 
    • Ideally, the following method should be used: Isopentane (methylbutane) should be cooled to below -100EC by placing it in a metal or plastic beaker and submerging it into liquid nitrogen.  If a low temperature thermometer is not available, you will know that the isopentane is approaching this temperature when it becomes white at the bottom of the container.
    • If this is not possible, the muscle should be frozen by the most rapid method with the lowest temperature possible.  We can usually overcome freeze artifact by thawing and refreezing the specimen.

  2. Assuming the two biopsies of sufficient size and quality have been taken, obtain a piece no larger than 2 X 1 X 1 mm. and place in glutaraldehyde for possible electron microscopy. Please use a screw capped or similar container.

  3. Shipping the specimen:
    • Local institutions:
      • The specimen is kept moist with very lighted saline-moistened gauze. The specimen should not be immersed in saline, or be allowed to dry.
      • It should be kept cool but not frozen.
    • Non-local institutions:
      • The muscle should be placed in a plastic bag and mailed in AT LEAST 10 POUNDS (5 KILOGRAMS) of dry ice.
      • The formalin and glutaraldehyde fixed tissue should be sent in leakproof screw top container. Some of the snap top containers that had been dipped in paraffin have had significant leakage by the time they reach us.

Dr. Hannes Vogel
Stanford University Medical Center
Neuropathology, Room R-241
300 Pasteur Drive
Palo Alto, CA 94305-5324

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