NEUROMUSCULAR PATHOLOGY CONSULTATION TECHNICAL INFORMATION

We have a comprehensive histochemistry panel for evaluation of neuromuscular diseases (1), and immunohistochemical panels for dystrophies (2) and inflammatory myopathies (3). In selected cases, we also perform ultrastructural analysis of muscle and peripheral nerve specimens at our electron microscope facility.

Panel 1: Histochemical Stains on Fresh Frozen Skeletal Muscle Specimen

Gomori Modified Trichrome
Succinyl Dehydrogenase
NADH
Cytochrome Oxidase
PAS with Diastase
Oil-Red-O
Acid Phosphatase
Alkaline Phosphatase
Non-Specific Esterase
ATPase (pH 9.4 and 4.5)
Congo Red

Panel 2: Immunohistochemical Stains for Dystrophies

Spectrin
Dystrophin (N-terminal, C-terminal, Rod-domain)
Alpha, beta, delta, and gamma- Sarcoglycans
Beta-Dystroglycan
Utrophin
Merosin
Merosin-M
Emerin
Caveolin-3
Dysferlin
Collagen IV, Collagen VI
Laminin beta-1
Lamin A/C

Panel 3: Inflammatory Myopathy Panel

CD3
CD20
CD8
CD4
CD68
Ubiquitin
MHC-Class 1
C5b-9
Congo Red with polarization and fluorescence microscopy

Panel 4: Metabolic Myopathy Panel

Modified SDH/COX double stain
Myophosphorylase
Myoadenylate Deaminase
PAS on plastic-embedded muscle sample
Electron microscopy for abnormal mitochondria, excess glycogen and lipid

Peripheral Nerve

All peripheral nerve samples are fixed in glutaraldehyde for electron microscopy, as well as in paraffin for light microscopy. For suspected cases of monoclonal gammopathy, fresh tissue is frozen for immunofluorescence studies.

Skin Biopsy For Intraepidermal Nerve Fiber Density Evaluation

For suspected cases of small-fiber neuropathy, PGP 9.5 immunostain is employed on skin biopsies for intraepidermal nerve fiber density evaluation.  This analysis requires a special fixative.  Please see protocol below.

PROTOCOLS FOR MUSCLE AND NERVE BIOPSIES

GENERAL

Correct interpretation of muscle and nerve biopsies is strongly dependent on the clinical setting. It is therefore imperative that pertinent clinical information, including medications and the name of the physician responsible for the patient, accompanies the specimen. This may be either faxed (503-494-6787) the day the specimen is shipped, or preferably e-mailed to NP@ohsu.edu. E-mailed information may be sent in advance as long as the approximate date of surgery is indicated.

MUSCLE

If a specific muscle is not dictated by the clinical presentation, the following muscles are preferred (in order of preference):

Quadriceps
Biceps
Deltoid

Adequate Size:

1 cm x 0.5 cm x 0.5 cm

Needle biopsies are acceptable but not recommended.
Avoid end stage muscle.
No infiltration with local anesthetic.
No clamps, or tying to tongue blade.
Do not freeze specimen.

Preparation for shipment:

Wrap specimen loosely in saline-moistened gauze or Telfa. Do not immerse in saline.
Place wrapped specimen into dry container e.g. specimen cup, and seal.
Surround container with wet ice. Do not ship on dry ice.
Pack in a Styrofoam container and Ship via courier, cab, FedEx or air.

If the specimen cannot be delivered to OHSU by 4:00 pm, the following procedure can be followed:

The specimen should be retained at the sending lab until the following OHSU workday. Specimens should be kept on saline-moistened (not soaked) gauze, in a dry closed container, in the refrigerator. If the following day is a Saturday or a holiday, then the specimen should be held (even over the weekend if necessary), until the next available OHSU workday. The next morning, OHSU should be contacted prior to shipping, and then the specimen shipped in the morning on moistened gauze, in a dry container, on wet ice, to Lab Central at OHSU. This is not the recommended treatment for optimal preservation of the specimen. Consideration should be given to rescheduling the biopsy to allow coordination between the sending facility and OHSU.

Nerve

Preferred length: 1.5 cm Avoid stretching or crushing nerve. Keeping the fresh nerve straight, place it on a narrow strip of filter paper, cut slightly longer than nerve. Immerse in 10% neutral buffered formalin (NBF). The nerve will adhere to the paper and this will maintain the orientation of the fibers. Exception: if the clinician is suspecting paraproteinemia-associated neuropathy, send two samples: one fresh or in transport medium for immunofluorescence and another fixed in 10% NBF for light and electron microscopy.  When we receive the nerve specimen, we will transfer a portion of the biopsy to glutaraldehyde.

Skin Biopsy For Small Fiber Neuropathy

Suspected cases of small fiber neuropathy undergo a standard 3mm punch biopsy of the skin in distal and proximal extremity locations. These tissues are processed with special tissue techniques and labeled with a PGP 9.5 antibody to quantify the intraepidermal nerve fibers with the light microscope. The results are expressed as a numerical figure as the number of nerve fibers per mm of epidermis and compared with published normal values.  These biopsies require a special fixative which we will provide.  Fixation time must be carefully controlled and special processing is needed, so we request that these biopsies be performed Monday-Thursday only.

Please contact immunohistochemistry for transport kit and instructions at (503) 494-5755, at least one day before the biopsy.

Shipping

Arrival at OHSU should be no later than 4:00 pm.

Notify Florence Leomiti at (503) 494-6781 9:00 am to 4:30 pm before sending the specimen with expected delivery time and additional pertinent information, e.g., flight number and arrival time, air waybill number, FedEx tracking number.

Cab or Courier:

Surgical Pathology
Room 3020, 5th Floor
Dillehunt Hall
OHSU
3181 SW Sam Jackson Park Road
Portland, OR, 97239

Notify Flo Leomiti, (503) 494-6781, upon arrival.

FedEx:

Marjorie Grafe, MD, PhD
Neuromuscular Lab L471
OHSU
3181 Sam Jackson Park Road
Portland, OR 97239-3098
Attn: Flo Leomiti, (503) 494-6781

Air:

Marjorie Grafe, MD, PhD
Neuromuscular Lab L471
OHSU
3181 Sam Jackson Park Road
Portland, OR 97239-3098
Attn: Flo Leomiti, (503) 494-6781

Shipping kits are available containing everything necessary to ship a muscle or nerve biopsy to OHSU. If you are interested in receiving a shipping kit please contact Florence Leomiti at (503) 494-6781 at least three days in advance of the collection of the specimen so a kit can be sent overnight to you.

ONCOLOGIC NEUROPATHOLOGY – CONSULTATION INFORMATION

Consultations from outside institutions are accepted in the form of glass slides and tissue blocks. Accompanying clinical information (age, site, duration, symptoms, known systemic cancer) and radiological data is essential for accurate diagnostic assessment.

Our immunohistochemical armamentarium includes the following:

• Glioma Panel: GFAP, Vimentin, p53, Ki-67, Phosphohistone H3, olig2, IDH1 mutation-specific antibody
• Pituitary Adenoma Panel: hGH, Prolactin, ACTH, SF-1, CAM 5.2, Reticulin
• Glioneuronal Tumors: Synaptophysin, Neurofilament protein, Neu-N, Chromogranin, CD34, nestin
• ATRT vs. Medulloblastoma: INI-1

In collaboration with the Knight Diagnostic Laboratories, we also offer fluorescent in situ hybridization (FISH) testing and DNA analysis on paraffin-embedded tissue sections for the following:

• Chromosome arms 1p and 19q
• EGFR amplification
• PTEN (10q) deletion and monosomy 10
• BRAF fusion
• BRAF V600E mutation
• MGMT promoter hypermethylation
• IDH1/2 mutations
• H3F3A mutation