Clinical Neurochemistry
Laboratory
|
Clinical Neurochemistry
Laboratory
|
|
|
Procedures for Sampling and Shipping of Samples ____________________________________________________________________________ TABLE OF CONTENTS
____________________________________________________________________________ PLASMA METANEPHRINES AND CATECHOLS
____________________________________________________________________________ 1. INTERFERENCES. Acetaminophen interferes with the plasma metanephrine assay. It is therefore essential that the patient must not have taken acetaminophen in any form (e.g., Tylenol, Excedrin, combined cold medications) for at least 5 days before the sample is drawn. Several medications contain or are catechols and may interfere with the results of the catechol assay. These include alpha-methyldopa, alpha-methyl-para-tyrosine, isoproterenol, dobutamine, and carbidopa (such as in Sinemet™). In addition, dihydrocaffeic acid, a catechol metabolite of caffeic acid, can occur at high plasma concentrations in people who have recently ingested caffeinated or even decaffeinated coffee. For these reasons, we ask that all medications the patient is taking be listed on the accompanying information sheet. It is best if the sample be obtained in the morning after an overnight fast (water and non-caffeinated soft drinks are permissible). 2. BLOOD COLLECTION. The patient should be in the supine position for at least 15 minutes before and during collection of the blood sample. To avoid the acute effects of the stress of venipuncture on plasma catechols and metanephrines it is essential that an indwelling i.v. be placed in a vein at least 15 minutes before the blood sample is drawn. The sample may be drawn through an indwelling butterfly i.v. set or through an i.v. cannula (normal saline to keep the line patent), via a 3-way stopcock and Vacutainer adapter into a waste tube (saline void) and then into a chilled evacuated glass collection tube containing heparin in any form as an anticoagulant (e.g., a green top Vacutainer tube). It is preferable, but not essential, to draw the sample without a tourniquet. 3. SAMPLE PROCESSING. The blood sample (10 ml) should be stored on ice until centrifuged (preferably at 4oC) to separate the plasma within 2 hours of blood collection. The plasma should be separated into two aliquots—one for catechols, one for metanephrines—with each aliquot at least 2 ml) in plastic tubes clearly marked with the date and patient ID and frozen immediately (e.g., on dry ice). The sample should be stored at -70oC or colder. Catechols in plasma stored at -20oC are not stable for extended periods, and so storage at -20oC should be limited to 2 weeks at most. 4. SPECIMEN SHIPPING. It is mandatory that personnel to receive the shipped sample be notified by phone, fax, or e-mail (see below) about the shipment and agree in advance to receive the shipment when it arrives. Samples of plasma should be shipped frozen (e.g., on dry ice in a styrofoam container) to the NIH. Please send Monday to Thursday, to ensure the samples arrive on a US Government business day between 9 AM and 5 PM. Please also ensure that shipments are labeled as containing dry ice (i.e., there should be a dry ice sticker).
5. ACCOMPANYING DATA. All samples should be shipped accompanied by the attached Patient Information Sheet. All portions of the top part of the patient information sheet must be filled out to satisfy CLIA guidelines and for test results to be sent back to the ordering physician. We also ask that a signed copy of the Patient Consent Form be included with all samples. 6. RESULTS. Assay results will be made available, but this may take up to 6 weeks after delivery of the sample (urgent requests may be processed sooner after consultation). Since the service is free (excluding the costs of shipping), the patient must not be billed for these determinations. INSTRUCTIONS FOR BLOOD SAMPLING AND SHIPPING (Detailed Instructions for Samples for Catechols From International Destinations) ________________________________________________________________________
2. Blood sampling Posture and stress. Catecholamines are sensitive to posture and any kind of stress. Therefore, samples should be routinely collected after the subject has rested for at least 15 minutes in one of several positions (e.g., supine of semi-supine) and preferably collected using an indwelling cannula or i.v. line inserted at least 15 minutes before sampling. Arterialization of blood samples. Most easily interpretable results are obtained with arterial blood samples, but this is often impractical in clinical settings. The preference for arterial over venous samples is secondary to the fact that up to 80% of circulating catecholamines are extracted during passage from the arterial to the venous circulation. This proportion varies depending on blood flow and vasoconstriction in the limb (this in turn largely dependent on ambient temperature). This means that levels of adrenaline in venous plasma are much lower than those in arterial plasma and much harder to detect; they are also dependent on local removal by the limbon in addition to release from the adrenal gland. Levels of noradrenaline reflect both removal by the limb and local release from sympathetic nerves of the limb and do not provide an accurate reflection of overall sympathetic outflow. Arterialization of venous blood provides a technique that helps overcome the above limitations of catecholamine determinations in venous samples. The technique simply involves gentle warming of the limb before standard collection of venous samples. This substantially increases blood flow so that extraction fractions of catecholamines (i.e., % of arterial catecholamine removed during passage through to the venous circulation) are reduced and the effect of local noradrenaline release is diluted out by the larger blood flow. Concentrations of adrenaline are also easier to detect and blood samples may be collected more easily. Diet. A recent meal can have a major influence on some catechols (e.g., plasma DOPA, DOPAC and sulfate conjugated dopamine). Therefore, the influence of diet should be controlled for wherever possible. 3. Sample volume At least 1.2 mL of plasma is preferred, ideally provided as duplicate samples to allow for repeat assays if required. Please note, however, the assay can be conducted on a minimum of 0.2 mL plasma (requires 0.45 - 0.5 mL blood). This should still enable determination of plasma concentrations of free adrenaline (normally <30 pg/mL in venous plasma), but concentrations of free dopamine are likely to be below the detection limits of the HPLC-EC assay (dopamine normally < 10 pg/mL). 4. Sample preparation Once collected, blood samples should be immediately transferred into heparinized tubes and stored on ice until centrifuged (ideally with 3 hr of collection) at 4 oC for 15 min to separate the plasma. If samples are collected in eppendorf tubes a microcentrifuge may be used, in which case the high spin rates allow separation of plasma from blood cells with 2 minutes. Although a refrigerated microcentrifuge (or one used in a cold room) remains preferable, use of reduced spin times makes it possible to centrifuge at room temperature.Once separated, plasma should be immediately transferred into small polypropylene tubes (glass and polystyrene tubes tend to crack at low temperatures or during subsequent centrifugations) and frozen on dry ice. Samples must be stored in a freezer at -70 oC or less. All tubes containing plasma should be appropriately labelled (use a code system and omit patient names) using an indelible marker.
5. Shipping of samples The NIH has agents located at Washington Dulles Airport (Wall shipping Inc.) who efficiently facilitate transfer, through US Customs, of shipments arriving at Dulles airport as the port of entry. These agents also organize transfer of the shipment from the Airport to the NIH. Therefore, shipments from Europe to the NIH must arrive at Washington Dulles as the port of entry. Shipments are best made using an airline carrier rather than a specialized international shipping company (also considerably cheaper). Samples should be placed in a sealed styrofoam box with sufficient dry ice (preferably block ice with crushed ice to fill the gaps) to last 4 days (normally samples sent from Europe arrive within 2 1/2 days - the extra dry ice helps avoid problems in case of unexpected delay). Samples should always be sent early in the week (Monday or Tuesday). Samples should be labeled as HUMAN PLASMA: NON-INFECTIOUS. A low monetary value must be assigned to the samples to facilitate transfer through customs. Finally, before the samples are sent, confirmation must be received by phone, fax, or e-mail (see below) that appropriate personnel will be available to receive the shipment. The airline, flight number, date and time of departure and expected time and date of arrival at Dulles must also be provided along with a faxed copy of the airway bill (or at least the airway bill number). Provision of this information makes it possible to notify our agents at Dulles of the shipments expected arrival time and allows our agents time to prepare the necessary documentation for expedited customs clearance. The shipment should be addressed to:
INSTRUCTIONS FOR BLOOD SAMPLING AND TRANSPORT OF PLASMA SAMPLES FOR ANALYSES OF CATECHOL OR METANEPHRINE CONCENTRATIONS (Detailed Instructions for Samples collected by Phlebotomy within NIH) ____________________________________________________________________________ Phlebotomy Procedures for Plasma Catechols and Metanephrines NIH Clinical Protocol 89-C-0086 Arlene Berman, RN (Beeper 104-5167-7-#), ACRF 3 Clinic Graeme Eisenhofer, Clinical Neuroscience Branch, NINDS (6-8925) ____________________________________________________________________________
____________________________________________________________________________ TIME IS OF THE ESSENCE IN APPROPRIATE HANDLING OF THE SPECIMEN. THE SPECIMEN MUST BE IN ICE UPON RECEIPT, OR ELSE IT WILL BE REJECTED. ____________________________________________________________________________
Patient's name: _______________________________________ NIH ID: _________________ Family Name, First Name, Middle Name (for NIH patients only) Birth date: ______________ Gender:_______ Date/time blood drawn: __________________ mth/day/yr m/f mth/day/yr - hr
Ordering Physician's Name (print legibly): ____________________________________________ Physician's phone no: _______________________ FAX no: _______________________
Physician's address: _____________________________________________________________ _____________________________________________________________ Tests requested (check each test requested): o Catechols o MetanephrinesReason for Test: _________________________________________________________________
Information below is not essential for report to be returned to ordering physician Medications (prescribed or over-the-counter) that the patient has taken in the last week: (Note: for tests of metanephrines patient must not have taken Acetaminophen or Tylenol for at least 5 days, since the drug interferes with test results)_____________________________________ _____________________________________ _____________________________________ _____________________________________ _____________________________________ _____________________________________ Sampling conditions (v) to confirm: Through i.v.? __ Supine = 15 min? ___ Fasted overnight? ___ Caffeinated or decaffeinated coffee in the past 12 hours? ___ Stored at -70 oC? ___
Vital signs: BP, P supine: _________________ BP, P upright 5 minutes: _________________ Please include any other pertinent clinical data (including results of specialized tests such as CT, MRI, or MIBG scans; urine catecholamines, metanephrines, or VMA; clonidine suppression test; tilt table test; Valsalva — Attach copies of lab reports or a patient history if appropriate). ______________________________________________________________________________
|
