Flow Cytometry Laboratory

Peripheral blood samples should be drawn in a sodium heparin or EDTA tube and kept at room temperature (22-28°C) until testing. Between one and eight milliliters of blood is required, depending upon the leukocyte count; a 5-ml sample is normally adequate. Samples should not be refrigerated; they must be stored at room temperature. Samples must be delivered to this laboratory as soon as possible or within 4 hours after collection, using transporters, couriers, or overnight delivery services. Immunology peripheral blood samples received more than 72 hours after collection will be rejected, as well as clotted, excessively hemolyzed, refrigerated or frozen specimens. Samples drawn in lithium heparin or ACD are not acceptable. CBC results drawn at the time of the flow specimen should be sent with the requisition.

Bone marrow samples should be drawn in a sodium heparinized syringe and transferred to a green top (sodium heparin) tube for transport. The sample must be kept at room temperature until testing. EDTA tubes are acceptable, but they must be less than 4 hours old when received. ACD and lithium heparin are not acceptable. Between 1 and 3 ml of bone marrow is required. Stability, storage, and transport of these samples are the same as that for peripheral blood. CBC results drawn at the time of the flow specimen should be sent with the requisition.

Lymph node or fresh biopsy tissue is acceptable. To ensure that adequate cells are available for flow cytometry, the tissue should be ≥ 1-2 cm square. Smaller samples are acceptable but may limit the number of markers that are able to be performed. The sample should be placed immediately in cold (2-8°C) RPMI media or on cold saline-soaked gauze and transported immediately on ice (not frozen) to the laboratory for testing. Receipt should be within 4 hours. If longer transport or overnight storage is required, the tissue should be kept in cold RPMI media and transported in the RPMI media on wet ice. Samples that have been exposed to fixatives (such as formalin or alcohol) are unacceptable for testing, as are samples that have been previously frozen or are frozen when received.

Body fluids (pleural, ascites, pericardial, cerebral spinal or CSF) and WASHINGS (fine needle aspirates or lung lavages) are acceptable for flow cytometric analysis if the cell number is sufficient. The minimal volume of sample required is 3 mls; however, depending upon cell number, this volume may not be sufficient for flow cytometric analysis. If the specimen is bloody, sodium heparin may be added to the specimen at collection. The sample should be kept at room temperature and delivered immediately to the laboratory. Receipt should be within 4 hours.

Processed samples of hematopoietic progenitor cell (HPC) harvests (apheresis, bone marrow) are acceptable for flow cytometric analysis of CD34+ stem cells. Samples should be sent at room temperature within 4 hours after processing.