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            SAMPLE COLLECTION MANUAL OF LABORATORY SERVICES
                    SIR T HOSPITAL, BHAVNAGAR 

Issue No: 04

Issue Date: 01/11/2019

Copy No: 01

Holder’s Name: Laboratory Director

CONTROLLED COPY

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II. CONTACT INFORMATION

Postal Address: Sir T General Hospital, Jail Road, Bhavnagar, Gujarat Pin-364001

Routine Day Enquiries

Contact details of the sections are:

                        IV  RELEASE AUTHORIZATION

The sample collection Manual is prepared as per ISO 15189:2012. This is the property of Sir T Hospital & Government Medical College Bhavnagar and issued under the authority of the undersigned.

                               V. AMENDMENT   RECORD   SHEET

                               VI. DISTRIBUTION LIST

The following are the authorized holders of the controlled soft copy only of sample collection manual.

                                VII  TABLE OF CONTENTS

Purpose: Laboratory testing provides information about a patient's health to assist Physicians in diagnostic and therapeutic decisions. Specimen Integrity is dependent on accurate pre-analytical processes to include patient preparation, specimen collection, handling, and transportation. Improper collection and handling of samples can give erroneous results and compromise the care of the patient. These guidelines cover some of the key steps in handling blood samples to provide optimal specimens for testing.

Principle: There are multiple Pre-Analytical factors associated with the handling and processing of laboratory specimens that can lead to test result inaccuracy. Strict adherence to all phases of proper collection and processing is essential for accurate test results.

Pre-Analytical Factors:

1. Specimen Collection:

• Improper Patient Identification
• Incorrect Order of Draw
• Incorrect Tube Selection
• Traumatic draws leading to hemolysis
• Inadequate mixing or insufficient sample

2. Specimen Handling/Processing:

• Serum tubes not thoroughly clotted before centrifugation
• Delay in Centrifugation
• Storing specimens in incorrect temperatures

3. Specimen Transportation

• Frozen specimens thawing during transport
• Unspun specimens transported >2 hours from collection

                       VIII Documentary Procedure of Patient identification

Purpose: To ensure accurate patient identification before phlebotomy to prevent mislabeling of specimens and ensure patient safety.

Responsibility: Phlebotomy personnel

Procedure: The phlebotomist's role requires a Greet the patient and identify yourself and indicate the procedure that will take place. Prior to any patient procedure, proper identification is a priority. Proper identification is a three-step process.

• Ask the patient to state his/her full name, Age, Mobile number
• Verify the details on the test requisition
• For unconscious patients & Pediatrics patients ask the caregiver to identify the patient.

                          Documentary Procedure of Patient Preparation

• Ask for Fasting status when relevant.
• History of medication
• For ambulatory patients blood should be collected in Seated position.
• For bedridden patients should be collected in supine position.
• Privacy & Confidentiality should be maintained.
• At no time should venipuncture be performed on a standing patient
• Arms should be extended in a straight line from the shoulder to the wrist
• Arm with an Previously inserted intravenous line should be avoided, as should an arm with extensive scarring or a hematoma.
• If fluid is being infused intravenously into a limb, the fluid should be shut off for 3 minutes.
• Cannula/fistula/heparin lock - Blood should not be drawn from an arm with a fistula or cannula without consulting the attending physician.
• Edematous extremities - tissue fluid accumulation alters test results.
• Cannula/fistula/heparin lock - Blood should not be drawn from an arm with a fistula or cannula without consulting the attending physician. Edematous extremities - tissue fluid accumulation alters test results.
• If h/o mastectomy then arm veins on that side of the body should not be used, because of lymphostasis. if double mastectomies, blood should be drawn from the arm of the side on which the first procedure was performed. If the surgery was done within 6 months on both sides, a vein on the back of the hand or at the ankle should be used.
• Children are likely to make unexpected movements, and assistance in holding them still is often desirable.

                           Documentary Procedure of Venipuncture
                           

Collection of blood from the vein is called venipuncture. Also referred to as phlebotomy. Assemble the necessary materials and equipment:

Wear the Personal Protective Equipments:

• A phlebotomist should dress in personal protective equipment (PPE)
• Must wear a Gloves

Selection of Site of Puncture:

• Veins of the forearm are preferred;veins in the antecubital fossa of the arm are the preferred sites.
• Wrist or ankle can also be used although these are less desirable and should be avoided in people with diabetes and other Individuals with poor circulation.
• The three main veins in the forearm

1. Medial Cubital- First choice. Anchored and easy to penetrate
2. Cephalic - On the outside surface, Well anchored
3. Basilic - Not well anchored, tends to roll, painful

• Palpate and trace the path of veins with the index finger. Arteries pulsate, are most elastic, and have a thick wall. Thrombosed veins lack resilience, feel cord-like, and roll easily.

Preparation of Site:

• Pre-packaged alcohol swab or Gauze pad saturated with 70% isopropanol or Benzalkonium chloride solution (Zephiran chloride solution, 1 : 750)
• Cleaning of the puncture site should be done with a circular motion and from the site outward.
• The skin should be allowed to dry in the air.
• No alcohol or cleanser should remain on the skin because traces may cause hemolysis.
• Do not re-touch the cleaned area

Venous Occlusion:

• Blood pressure cuff (60 mm Hg)or a tourniquet is applied 4 to 6 inches (10 to 15 cm) above the intended puncture site (distance for adults)

Procedure for collection using Evacuated system:

• The short end of the double-pointed needle enters the holder and pushes the tube forward until the top of the stopper meets the guide mark on the holder.

• The point of the needle will thus be embedded in the stopper without puncturing it and losing the vacuum in the tube.

• Insert the needle properly into the vein, the index finger is placed alongside the hub of the needle with the bevel facing up. The needle should be pointing in the same direction as the vein.
• Then the point of the needle is advanced 0.5-1.0cm into the subcutaneous tissue (at an angle of 450) and is pushed forward at a lesser angle to pierce the vein wall.
• When the needle is properly in the vein, the vacuum tube is pushed into the needle holder all the way so that the blood flows into the tube under vacuum.
• The tourniquet should be released the moment blood starts entering the vacuum tube otherwise, some hemoconcentration will develop after one minute of venous stasis.
• After drawing the required blood sample, apply a ball of cotton to the puncture site and gently withdraw the needle.
• Instruct the patient to press on the cotton
• Remove the tube from the vacutainer holder and if the tube is with anticoagulant, gently invert several times
• Label the tube information required by the hospital (before the patient leaves the collection area) Re-inspect the venipuncture site to ascertain that the bleeding has stopped. Do not let the patient go until the bleeding stops.
• Blood collected into a tube containing one additive should never be transferred into other tubes, because the first additive may interfere with tests for which a different additive is specified.

Order Of Draw: Specific order for Blood collection tubes is followed to avoid cross-contamination of additives between tubes. The recommended order of draw is:

1. Plain (Red top)
2. SST (red-gray or gold top). Contains a gel separator and clot activator.
3. Sodium heparin (dark green top)
4. PST (light green top). Contains lithium heparin anticoagulant and a gel separator.
5. EDTA (lavender top)
6. ACDA or ACDB (pale yellow top). Contains acid citrate dextrose.
7. Oxalate/fluoride (light gray top)

NOTE: Tubes with additives must be thoroughly mixed, otherwise erroneous test results may be obtained when the blood is not thoroughly mixed with the additive.

                Venepuncture technique using needle and syringe

• Remove the syringe and the needle from its protective wrapper.
• Assemble them allowing the cap to remain covering theneedle until use.
• Check to make sure:

1. The needle is sharp
2. The syringe moves smoothly
3. There is no air left in the barrel

• Insert the needle properly into the vein, the index finger is placed alongside the hub of the needle with the bevel facing up. The needle should be pointing in the same direction as the vein.
• Then the point of the needle is advanced 0.5-1.0cm into the subcutaneous tissue (at an angle of 450) and is pushed forward at a lesser angle to pierce the vein wall.
• When the needle is properly in the vein plunger is drown back to create suction pressure to draw the blood.

Venepuncture in Children A syringe should be the tuberculin type or should have a 3- mL capacity, except when a large volume of blood is required for analysis

                   Documentary  Procedure of Capillary blood collection

Also referred to as micro blood samples collection/dermal puncture

Indications:

• In infants - Preferred when only small quantities of blood are required i.e.for hemoglobin quantitation, for WBC and RBC counts and for blood smear preparation
• In cases of severe burns
• In extreme obesity where locating the veins could be a problem
• In patient whose arm veins are being used for intravenous medication

Sites of Puncture:

• Adults and children: Palmer surface of the tip of the ring or middle finger or free margin of the ear lobe
• Infants: plantar surface of the big toe or the heel.
• Edematous, congested and cyanotic sites should not be punctured.
• Cold sites should not be punctured as samples collected from cold sites give falsely high results of hemoglobin and cell counts.
• Site should be massaged until it is warm.

Materials Required: Cotton or gauze pads 70% alcohol or other skin antiseptic Sterile disposable lancet (Automatic lancet) Blood collection tubes

Technique:

• Rub the site vigorously with cotton or a gauze pad moistened with 70% alcohol To remove dirt and epithelial debris and To increase blood circulation in the area If the heel is to be punctured, it should first be warmed by immersion in warm water or applying a hot towel compress. Otherwise values significantly higher than those in venous blood may be obtained.
• After the skin has dried, make a puncture 2-3 mm deep with a sterile lancet.
• A rapid and firm puncture should be made with control of the depth.
• A deep puncture is no more painful than a superficial one and makes repeated punctures unnecessary.
• The first drop of blood which contains tissue juices should be wiped away.
• The site should not be squeezed or pressed to get blood since this dilutes it with fluid from the tissues.
• a freely flowing blood should be taken or a moderate pressure some distance above the puncture site is allowable.
• Stop the blood flow by applying slight pressure at the site preferably with a gauze pad.
• Cotton balls are not recommended, because they stick to the site and can initiate bleeding.

                      Factors  interfere with test results

Hemolysis can interfere with many test results.

• Mix tubes with anticoagulant additives gently 5-10 times
• Avoid drawing blood from a hematoma
• Avoid drawing the plunger back too forcefully, if using a needle and syringe, and avoid frothing of the sample
• Make sure the venipuncture site is dry
• Avoid probing, trauma

Indwelling Lines or Catheters:

• Potential source of test error
• Most lines are flushed with a solution of heparin to reduce the risk of thrombosis
• Discard a sample at least three times the volume of the line before a specimen is obtained for analysis

Hemoconcentration: An increased concentration of larger molecules and formed elements in the blood may be due to several factors:

• Prolonged tourniquet application (no more than 2 minutes)
• Massaging, squeezing, or probing a site
• Long-term IV therapy
• Sclerosed or occluded veins

Prolonged Tourniquet Application:

• Primary effect is hemoconcentration of non-filterable elements (i.e. proteins).
• Significant increases can be found in total protein, aspartate aminotransferase (AST), total lipids, cholesterol, iron.
• Affects packed cell volume and other cellular elements.

                  Documentary Procedure of Arterial Blood collection  

Arterial Blood collection

• Arterial blood sampling should only be performed by health workers who have demonstrated proficiency after formal training.
• The sample can be obtained either through a catheter placed in an artery, or by using a needle and syringe to puncture an artery.
• These syringes are pre-heparinized and handled to minimize air exposure that will alter the blood gas values.

Choice of site:

• First choice is radial artery
• Alternative sites for access are brachial or femoral arteries.

Equipment and supplies:

• Pre-heparinized syringe
• Needles (20, 23 and 25 gauge, of different lengths)
• Bandage to cover the puncture site after collection.
• Container with crushed ice for transportation of the sample to the laboratory (if the analysis is not done at the point of care).

Procedure for arterial blood sampling using radial artery:

• Approach the patient, introduce yourself and ask the patient to state their full name.
• Place the patient on their back, lying flat. Ask the nurse for assistance if the patient’s position needs to be altered to make them more comfortable.
• Locate the radial artery by Allen test or if necessary locate the site by palpation.
• Perform hand hygiene, clear off a bedside work area and prepare supplies. Put on an impervious gown or apron, and face protection, if exposure to blood is anticipated.
• Disinfect the sampling site on the patient with 70% alcohol and allow it to dry.
• If the needle and syringe are not preassembled, assemble the needle and heparinized syringe and pull the syringe plunger to the required fill level recommended by the local laboratory.
• Holding the syringe and needle like a dart, use the index finger to locate the pulse again, inform the patient that the skin is about to be pierced then insert the needle at a 45 degree angle, approximately 1 cm distal to (i.e. away from) the index finger, to avoid contaminating the area where the needle enters the skin.
• Advance the needle into the radial artery until a blood flashback appears, then allow the syringe to fill to the appropriate level. DO NOT pull back the syringe plunger.
• Withdraw the needle and syringe; place a clean, dry piece of gauze or cotton wool over the site and have the patient or an assistant apply firm pressure for sufficient time to stop the bleeding. Check whether bleeding has stopped after 2–3 minutes.
• Expel air bubbles, cap the syringe and roll the specimen between the hands to gently mix it. Cap the syringe to prevent contact between the arterial blood sample and the air, and to prevent leaking during transport to the laboratory on ice packs.
• Label the sample syringe.
• Dispose appropriately of all used material and personal protective equipment.
• Remove gloves and wash hands thoroughly with soap and water
• Check the patient site for bleeding (if necessary, apply additional pressure) and thank the patient.
• Transport the sample immediately to the laboratory, following laboratory handling procedures.

Caution and Care during arterial blood sampling:

• Arteriospasm may be prevented simply by helping the patient relax; this can be achieved, for example, by explaining the procedure
• Haematoma can be prevented by inserting the needle without puncturing the far side of the vessel and by applying pressure immediately after blood is drawn
• Nerve damage can be prevented by choosing an appropriate sampling site and avoiding redirection of the needle.
• Fainting or a vasovagal response can be prevented by ensuring that the patient is supine (lying down on their back) with feet elevated before beginning the blood draw.
• Other probems can include a drop in blood pressure, complaints of feeling faint, sweating or pallor that may precede a loss of consciousness.

Sampling errors:

* Presence of air in the sample * Collection of venous rather than arterial blood * An improper quantity of heparin in the syringe, or improper mixing after blood is drawn * A delay in specimen transportation.

Documentary Procedure of Urine Specimen collection

* Urine Specimen collection

Random: The normal composition of urine varies considerably during a 24-hour period. Most reference values are based on analysis of the first urine voided in the morning. This specimen is preferred because it has a more uniform volume and concentration, and its lower pH helps preserve the formed elements. Submit a first morning specimen whenever possible. Urine for pregnancy testing should be a first morning voiding, or a random specimen with a specific gravity of at least 1.010. Note the time of collection of the specimen on the test request form and on the label of the container. Submit urine for pregnancy testing in a plastic vial with no preservative. To reduce contamination, the specimen submitted for urinalysis should be a clean catch “midstream sample.” Urinalysis specimens must be submitted in a yellow capped urine transport tube containing preservative.

24-Hour Urine Collection: Proper collection and preservation of 24-hour urine specimens is essential for accurate test results. Patients are carefully instructed in the correct procedure.

A collection container and detailed instructions are given to the patient. If a urine preservative is required, it is ensured that the designated preservative is added in the urine collection container at the start of the collection. The patient is instructed that the preservative may be toxic and caustic and that it should not be spilled or discarded. Record any medication that the patient is receiving on the test request form. Mix the urine well, measure the volume of the 24-hour collection and record volume on both the test request form and the transport vial. Transfer the required volume into a plain urine transport vial. Add any additional required preservative and mix well. Do not send the entire urine collection.

Note: For those analyses requiring the addition of 6N HCI or other preservatives, add the preservative at the start of collection. Have the patient collect each specimen in a smaller container and carefully pour the urine into the 24-hour container to avoid any possible acid burns to the patient. Be sure to mix urine thoroughly before removing the aliquot.

Instructions to the Patient:

1. Avoid alcoholic beverages and vitamins for at least 24 hours before starting to collect urine, and during the collection period. Do not discontinue medications unless instructed to do so by your physician. Inform the laboratory which medications you are taking.
2. Do not exceed your normal intake of liquids or change your dietary habits during the day before and the day of your collection unless your physician gives you specific instructions to do so.
3. Empty bladder (void) into the toilet on the morning of the collection day. Do not include the first urine specimen of the day.
4. Collect all subsequent urine voided for the next 24 hours and add to the container provided by the laboratory. The last sample collected should be the first specimen of the following morning at the same time as the previous morning's first voiding.
5. Keep the urine in a cool place. Refrigerate if possible.
6. Deliver to the referring healthcare provider promptly.
7. Preservative may have been added to this container. This may be caustic. Please be careful not to spill any of the preservative.

                    Documentary Procedure of Safety and Infection Control                                   

* Because of contacts with sick patients and their specimens, it is important to follow safety and infection control procedures.

Self Protection: Standard Precautions:

1. Wear gloves and a lab coat or gown when handling blood/body fluids.
2. Change gloves after each patient or when contaminated.
3. Wash hands frequently.

• Dispose off items in appropriate containers. Dispose of f needles immediately upon removal from the patient's vein. Do not bend, break, recap, or resheath needles to avoid accidental needle puncture or splashing of contents.
• Clean up any blood spills with a disinfectant such as freshly made 10% bleach.

If you prick yourself with a contaminated needle:

• Remove your gloves and dispose them off properly.
• Squeeze puncture site to promote bleeding.
• Wash the area well with soap and water.
• Record the patient's name and ID number.

Follow the institution's guidelines regarding treatment & follow-up. Note: The use of prophylactic zidovudine following blood exposure to HIV has shown effectiveness (about 79%) in preventing seroconversion.

Patient Precaution:

• Place blood collection equipment away from patients, especially children and psychiatric patients.
• Practice hygiene for the patient's protection. When wearing gloves, change them between each patient and wash your hands frequently. Always wear a clean lab coat or gown.
• Sterile venipuncture.