SENDING SPECIMENS

• Specimen Requirements & Recommended Tissue Handling
  • Flow Cytometry
  • Fluorescence in situ Hybridization (FISH)
  • Immunohistochemistry (IHC)
  • Immunofluorescence (IF) on skin and mucosal tissue
  • Polymerase Chain Reaction (PCR) for B and T cell Gene Rearrangements
• Detailed Specimen Shipping Instructions
  • General Requirements
  • Requisition Forms
  • Paraffin Blocks
  • Glass Slides
  • Slides or Tissues in Fixative
  • Tissues in IF Transport Media (Michel's, Zeus...)
  • Serum for IF
  • Tissue or Cytology Specimens in RPMI, Blood, Bone Marrow or Other Fresh Specimens
• Billing

Specimen Requirements & Recommended Tissue Handling

Call PhenoPath toll-free at 1-888-927-PHENO (888-927-4366) for questions, or to request shipping supplies.

For any case: DO NOT freeze specimen or transport medium (never ship on dry ice).

1. Label all specimen containers with the patient name and identifying number.
2. Place all specimen containers in leak proof biohazard bag with absorbent material. Use Ice Packs and a Styrofoam mailer when shipping.
3. Enclose the following in the package:
   • Completed PhenoPath Laboratories requisition form
   • Billing information
   • Clinical information
   • Mailer with specimen enclosed
4. Return to PhenoPath Laboratories via FedEx, courier, or other delivery method of choice.
   Please do not use US Mail Service. Free overnight delivery is available.

Saturday Delivery FedEx Billable Stamp
PhenoPath Laboratories can provide FedEx prepaid billable stamps for use with fresh specimens that need to arrive on Saturday (instead of Monday). These stamps MUST be used to ensure Saturday delivery. Please use these specific billable stamps only for fresh specimens that are sent on Friday to arrive on Saturday. If you have a specimen to send to PhenoPath Laboratories Monday thru Thursday, or on Friday for Monday delivery, please use the standard prepaid FedEx billable stamp (also supplied by PhenoPath Laboratories). Please let client services know if you are in need of additional billable stamps of either kind.

Flow Cytometry (for Leukemia/Lymphoma Diagnosis)

See more detailed flow cytometry instructions, including RPMI requirements.

1. Please include all relevant clinical data, including previous diagnoses, on a PhenoPath requisition form, along with copies of any current or previous flow cytometry or cytogenetic reports.
2. Specimen requirements:
• Peripheral blood: In general, 5 to 10 ml of anticoagulated peripheral blood is adequate, although for patients with marked peripheral leukopenia of less than 1000 white blood cells per µL, 10 to 20 ml should be considered. For flow cytometry alone, sodium heparin anticoagulation (green top tube) is preferred, but EDTA anticoagulation (lavender top tube) is also acceptable. When both flow cytometry and PCR studies are desired, separate heparin and EDTA-anticoagulated tubes, respectively, are preferred. Specimens should be sent with (previously frozen) cold packs.
• Bone marrow aspirate: 1 to 2 ml of anticoagulated bone marrow aspirate is adequate for high quality, cellular specimens with little hemodilution by peripheral blood. For flow cytometry alone, sodium heparin anticoagulation is preferred, but EDTA anticoagulation is also acceptable. When both flow cytometry and PCR studies are desired from a single specimen, EDTA anticoagulation is preferred. Specimens should be sent with (previously frozen) cold packs.
• Bone marrow biopsy: In patients with inaspirable bone marrow and insufficient neoplastic cells in the peripheral blood to enable a definitive diagnosis, it may be possible to obtain sufficient cells for flow cytometry from a bone marrow core biopsy submitted unfixed in tissue culture medium (RPMI-1640 with 10% fetal bovine serum [FBS]), which is supplied in plastic specimen vials by PhenoPath Laboratories. Specimens should be sent with (previously frozen) cold packs, and will be processed in the laboratory to maximize the release of cells from the bone core.
• Tissues: With the exception of fine needle aspirations, all tissues for flow cytometry, whether small needle core biopsies or relatively large portions of excisional biopsies or tumor resections, should be finely minced to maximize exfoliation of cells into the medium, and then transported in plastic vials containing RPMI-1640/10% FBS, which are supplied by PhenoPath. The tissue fragments should be completely submerged in the medium. Specimens should be sent with (previously frozen) cold packs, but under no circumstances should the cells be allowed to freeze, since this will destroy cell membranes.
• Cerebrospinal fluid (CSF): No less than 1 mL of non-traumatically-obtained CSF should be reserved for flow cytometry. Due to the relative absence of nutrients in CSF, specimens to be sent for flow cytometry should be combined with an equal or greater volume of RPMI-1640/10% FBS as soon as possible after the specimen is obtained. Containers should be closed tightly to prevent leakage, and sent with (previously frozen) cold packs.
• Note that the addition of RPMI-1640/10% FBS precludes determination of the absolute leukocyte count in vivo in this aliquot of CSF.
• Other body fluids, including bronchoalveolar lavage (BAL) fluids: For flow cytometry immunophenotyping, such fluids should be combined with an equal or greater volume of RPMI-1640/10% FBS prior to being sent with (previously frozen) cold packs.
Custom Flow Cytometry Panels: Customized antibody panels for clinical or research use can be designed in consultation with the laboratory director.

Fluorescence in situ Hybridization (FISH)

FISH tests offered at PhenoPath can be performed on formalin-fixed paraffin-embedded tissue, cytology material, fresh tissue, blood or bone marrow.

Immunohistochemistry (IHC)

Fixation: Virtually all IHC tests offered at PhenoPath Laboratories can be performed on formalin-fixed paraffin-embedded tissue, including all lymphoid marker studies (except CD52, which requires frozen tissue). While most antibodies may work on tissues fixed in non-formalin based fixatives, suboptimal tissue antigenic preservation may result and lead to false negative results; please contact the laboratory (1-800-92-PHENO; 1-800-927-4366) if you have questions about a specific fixative.

Decalcification: Virtually all antibodies can be applied to decalcified tissues; however, reactivity may be compromised in ways impossible to predict. As with all histological procedures, thorough fixation prior to decalcification is required. Checking the endpoint of decalcification to avoid over-treatment also improves the likelihood of adequate immunoreactivity.

Cut Sections: We generally prefer to receive the paraffin block, but if sending unstained sections, they must be on adhesive ("charged," "Plus," "APES," "silane," "poly-L-lysine"...) slides, preferably positioned at the center. Put one section per slide. Please call the laboratory if you are uncertain how many sections to send, or if your slides are acceptable (1-800-92-PHENO; 1-800-927-4366).

Cytology Specimens for IHC: Cytology specimens are acceptable for virtually all IHC tests; however, some antibodies may be less sensitive in the alcohol fixation typical of cytology specimens. These include TTF-1, ER, and PR, among others. Acceptable specimens include smears, touch preps and cytocentrifuge preps. In general, air-dried is preferred to in fixative. Cytology cell blocks are treated as any other formalin-fixed paraffin-embedded tissue.

Immunofluorescence (IF) on skin and mucosal tissue:

• Ship skin or other tissue specimens in IF Transport Media (see "Detailed Specimen
  Shipping Instructions."
• Frozen tissue is still the standard for immunofluorescence studies; however, formalin-fixed, paraffin-embedded tissue may be used as an alternative if frozen is not available. Sensitivity may be affected.
• Serum for IF: Serum may also be sent for IF studies. Minimum requirement is 0.5 mL of serum, but >1.0 mL is preferable.

Polymerase chain reaction (PCR) for B and T cell Gene Rearrangements

• Formalin-fixed paraffin-embedded tissue,
• Fresh (1 mL minimum in RPMI) or frozen tissue specimens,
• Body fluids (cytology material),
• Peripheral blood - EDTA anticoagulated (lavender top), or
• Bone marrow - EDTA anticoagulated (lavender top)

Detailed Specimen Shipping Instructions

Call Pheno Path toll-free at 1-888-92-PHENO (888-927-4366) for questions, or to request shipping supplies.

General Requirements

1. Send a requisition form completed according to the instructions (including ICD9 code and collection date). Preprinted requisition forms will be sent upon request, or can be downloaded below.
• Requisition Form - Hematopathology
• Requisition Form - HER2 Validation
• Requisition Form - All other pathology tests
• Material Request Form
2. Please include an H&E stained slide and a copy of the pathology report (even if preliminary) with all cases. This will aid our interpretation, and help avoid delays. If there is an area of particular interest that you would like us to review, please mark it on the H&E. We will return your H&E promptly upon completion of our studies, unless you indicate that we may keep it.
3. Standard overnight express shipping is provided by PhenoPath to ensure reliable and rapid delivery of specimens from your laboratory to ours. Prepaid air bills are provided. If sending perishable specimens on a Friday, please be sure to check the box for SATURDAY DELIVERY. If the specimen is non-perishable (blocks, slides, tissue in IF Transport Media or fixative), check the Standard Overnight box. DON'T mail specimens by regular U.S. Mail; always use a traceable means.
4. Days of Operation (Expanded, Fall 2005): PhenoPath Laboratories now accepts deliveries 6 days a week (Monday – Saturday). PhenoPath must be called on Friday to arrange for Saturday service. Saturday service available for flow cytometry specimens only.
5. Please contact us toll-free at (888) 92-PHENO (888-927-4366) with questions or to request preprinted requisition forms and shipping supplies (e.g., prepaid air bills, PhenoBoxes, etc.).

Paraffin Blocks

DO wrap blocks individually in gauze; DON'T wrap blocks in bubble wrap or plastic. DO place blocks in insulated styrofoam boxes provided by PhenoPath, and put in 1 to 2 small ice packs to prevent melting wax. Tape around edges of the styrofoam box to further insulate, and place into a PhenoBox.

Glass Slides

DO place glass slides in hard plastic, top loading slide containers, so that the slides do not rattle, and wrap in bubble wrap or pack in styrofoam peanuts; place all in a PhenoBox to prevent breakage. DON'T send slides in side-loading plastic containers or cardboard mailers.

Slides or Tissues in Fixative

DO properly label specimen with patient name and specimen ID. DO ensure that cap is tightly sealed. Shipping regulations require volumes of less than 50 mL. Put the vial containing fixative inside a secondary container (such as a plastic bag, or a larger vial) containing enough absorbent material to completely absorb any spill (e.g., paper towels, vermiculite, etc…). Place the secondary container into the PhenoBox for shipping. Slides and tissues in fixative may be shipped at room temperature.

Tissues in IF Transport Media (Michel's, Zeus…)

Skin or other tissue specimens for immunofluorescence should be placed into the IF Transport Media, which is provided by PhenoPath Laboratories, or you may use the following equivalent products: Michel's fixative, Zeus Transport Media. DON'T use RPMI, or flow cytometry transport media. Ensure the Transport Media pH is close to 7.0. DON'T use Transport Media for IF specimens if pH is lower than 6.0. DO properly label specimen with patient name and specimen ID. Tightly close the vial and place in the "Biohazard" bag provided with the vial. The paperwork should be folded into quarters, with the patient information on the inside, and placed in the pouch on the outside of the bag. The specimen may be sent at room temperature.

Serum for IF

The blood should be drawn into a clot tube (red top), spun down and the serum separated from the clot. The serum tube should be placed in a tube mailer, then a secondary container with enough absorbent material to completely absorb any spill (e.g., paper towels, vermiculite, etc…). Ship on wet or dry ice via overnight shipping.

Tissue or Cytology Specimens in RPMI, Blood, Bone Marrow, or Other Fresh Specimens

DO properly label specimen with patient name and specimen ID. DO ensure that cap is tightly sealed. The vial containing the specimen must be placed inside a secondary container (such as a plastic bag, or a larger vial) containing enough absorbent material to completely absorb any spill (e.g., paper towels, vermiculite, etc…). Place the secondary container into a styrofoam mailer on sufficient wet ice or cool packs for overnight shipping, and then into the cardboard overpack.

Billing

PhenoPath Laboratories provides billing directly to laboratories and other clients, as well as all third party payers, patients, and Medicare/Medicaid, as allowable by law. In order to ensure proper billing to patient and third-party payers, a copy of the patient’s registration file and ICD9 code and/or submitting pathology report are required. For details on our billing policies, please see our Billing Policies & Medicare Compliance page or contact our Billing Department at 206-374-1480, or toll free at 888-927-4366.