“PREPARATION, PROCESSING,

& HANDLING BLOOD & NON-

BLOOD SPECIMEN”

BMLS 1B

ALKUINO, REAH JAYZEL G.

I. Handling Blood Specimen
A. Mixing of Tubes
In mixing the tube, additive tubes must be mixed immediately after collection by inverting them
from three to eight times depending on the additive type. Hence, If tubes are not quickly and thoroughly
mixed, micro clots can form in anticoagulant tubes, and clotting may be incomplete in clot activator tubes.
In addition, Gentle inversion is essential, as vigorous mixing can cause hemolysis.
B. Transporting Specimen
The specimen must be transported stopper up to reduce agitation, allow serum tubes to clot
properly, and keep blood away from the stopper for safety reasons. Blood specimen tubes must always be
transported carefully to prevent breakage and protect specimen integrity. Also, Agitation or rough handling
can cause hemolysis and lead to platelet activation that can affect coagulation tests. As said by CLSI and
OSHA guidelines, blood specimen tubes must be transported to the laboratory in a plastic bag with a
biohazard logo, a liquid-tight closure, and a slip pocket for paperwork.
C. Delivery Time
The phlebotomist should arrange his or her scheduled based on the delivery time of the specimen
to prevent errors. Routine blood specimens should ideally be delivered to the laboratory within 45 minutes
of collection and centrifuged within 1 hour of arrival if serum or plasma is needed, while Stat specimens
must be transported, processed, and tested immediately. In this case, Prompt separation is essential to
minimize the effects of metabolic processes such as glycolysis.
D. Analyte Protection
Prewarming
Agglutination or Precipitation will occur if the specimen is being cooled below room temperature
(1-5C). It is typically transported in prewarmed tubes, heat blocks or wrapped in special warming material.
Chilling
In order to slow down metabolic processes that can negatively affect an analyte, it is required for
the specimen to undergo chilling. These specimens must be transported in crushed ice and water slurry.
Light Protection
If Bilirubin is exposed to light, decrease up to 50% in hour is observed. It must be wrapped in foil
carbon paper or collected in amber colored bottles for the analyte of the Specimens to protected from the
light.

II. Processing Blood Specimens

A. Processing safety
The use of Personal protective equipment including gloves, fluid resistant lab gowns, & protective
face gear by specimen processors are required for OSHA regulations. Hence, all procedures involving blood
must be performed in a manner that minimizes splashing, spraying, splattering, and generation of droplets.
B. Specimen Suitability
Chemistry specimens are most frequently rejected for hemolysis. It includes failure to follow
special timing or handling requirements, clotting in whole blood or plasma specimens, QNS (quantity not
sufficient), and collection in the wrong tube. Also, properly identified, quality specimens are required for
valid test results. Poor-quality or improperly identified specimens are normally rejected for testing, and new
specimens are requested.
C. Centrifugation
Centrifuge Machine is which centrifugal force is applied to separate solid matter from the liquid
suspension. Horizontal Head & Angle Head Centrifuge are the two main types of centrifuge. Also, if
performed on whole blood, a blood specimen should be centrifuged to separate the serum or plasma from
the cells. It includes Cytocentrifuge and Ultracentrifuge.
Horizontal Head Angle Head
 AKA Swinging Bucket  AKA Fixed Angle
 Stationary (V), Spinning (H)  Fixed
 Increased Air friction & Resistance, Max speed: 1650g  Decreased Air friction
 Remixing is Possible
 & Resistance, Max speed: 9000g
Rules on Centrifugation
1. Tubes placed in the cups of the rotor head must be equally balanced.
2. A centrifuge should never be operated until the top has been firmly fastened down, and the top
should never be opened until the rotor head has stopped completely
3. Do not walk away from the centrifuge until it has reached its rotational speed & no excessive
vibration is observed
4. When a tube breaks inside the centrifuge, immediately stop the spinning & unplug before opening
the lid.
5. If possible, Samples should not be recentrifuged.
D. Stopper Removal
The Stoppers must be enclosed with gauze or tissue to catch blood drops or aerosol that may be
released during removal and pulled straight up and off; they must not be popped off by a thumb roll
technique.
E. Aliquot Preparation
Aliquot is frequently created when multiple tests are ordered on a single specimen. It is prepared
by transferring a portion of the specimen into one or more tubes labeled with the same ID information as
the specimen tube. Also, it must be covered as soon as they are filled to prevent evaporation and protected
from heat and light. Most samples that are delayed for testing can be refrigerated at 4°C for up to 8 hours.

III. Urine
Urine samples include Random, 1st Morning, Midstream Clean Catch, 24-hr (timed) specimen,
Catheterized & Suprapubic Aspirate. Urine collection should be Midstream and it should be processed
within 2 hours.
A. Specimen Collection
In Specimen Collection, container should be clean, dry, leak-proof that should have a wide mouth
to facilitate collections from female patients and a wide flat bottom to prevent overturning. The capacity
should be 50 mL which allows 10-15 mL of the collected specimen.
In pediatric specimen, it should be collected from bags with adhesive. Large container with properly applied
screw top lids is required for the 24-hours urine collection. Sterile containers with secure closures are
needed for microbiological studies. In addition, Dark or Amber bottles are used for bilirubin, urobilinogen
and porphyrin analysis.
B. Specimen Labeling
The label of the specimen should have the following: Patient’s complete name, identification
number, date and time of collection, age and gender. The label should be attached to the container, not to
the lid, and should not become detached if the container is refrigerated or frozen. Also, it is accompanied
with a requisition form that matches the information on the specimen label.
Unacceptable Specimens are the following
 Unlabeled containers
 Nonmatching labels and requisition forms
 Specimens contaminated with feces or toilet papers
 Containers with contaminated exteriors
 Specimens of insufficient quantity
 Improperly transported specimens
C. Types of Urine Specimen
Random Urine
Samples are collected by patients while at the laboratory and are used primarily for routine
urinalysis. Patients is provided with a disposable urine container and directed to the bathroom facility.
1st Morning Urine
A first morning sample is the specimen of choice for urinalysis because it is more concentrated. It
is used to confirm tests previously done by Random Urine. Patients are provided with a container and
instructed to collect the sample immediately after arising and to return it to the laboratory within 2 hours or
refrigerate the sample.
Catheterized Urine
The Specimen is collected under sterile conditions by passing a hollow tube (catheter) through the
urethra into the bladder. It is commonly requested for bacterial culture. Also, it is requested to measure
functions in the individual kidneys.

Midstream Clean Catch

Midstream clean-catch samples are used for urine Aerobic Bacterial Culture. The Specimen is less
contaminated by epithelial cells and bacteria, therefore more representative of the actual urine than the
routinely voided specimen. Also, Mild antiseptic towelettes are recommended. Patients are instructed to
wash their hands before beginning the collection.
24-Hour Timed Urine
The sample is required for quantitative measurement of urine constituents. Patients are provided
with large plastic containers that may contain a preservative. The collection bottle must be larger than the
ones used in routine urine collection. In addition, to maintain the integrity of the samples, preservatives are
added.
24-Hour (Timed) Urine Sample Collection Procedure
1. Provide patient with written instructions and explain the collection procedure.
2. Issue the proper collection container and preservative.
3. Day 1: 7 a.m. Patient voids and discards specimen.
4. Patient writes the exact time on the sample label and places the label on the container.
5. Patient collects all urine for the next 24 hours.
6. Refrigerate the sample after adding each urine collection.
7. Patient should collect urine sample before bowel movement to avoid fecal contamination.
8. Continue to drink normal amounts of fluid throughout the collection time.
9. Day 2: 7 a.m. Patient voids and adds this urine to the previously collected urine.
10. Sample is transported to the laboratory in an insulated bag or portable cooler.
11. Upon arrival in the laboratory, the entire 24-hour sample is thoroughly mixed, and the volume is
accurately measure and recorded.
12. An aliquot is saved for testing and additional or repeat testing. The remaining urine is discarded.

Suprapubic Aspirate
A suprapubic sample is collected by external introduction of a needle through the abdomen into the
bladder. Provides a sample for Anaerobic bacterial culture that is completely free of extraneous
contamination. Also, Specimen may be used for cytologic examination.
D. Physical Examination of Urine
Urine color
It is a rough indicator of the degree of hydration

NORMAL ABNORMAL

Colorless Red

Pale yellow Red brown

(Diluted Urine)
Dark yellow (concentrated
Urine)
HOW TO EXAMINE URINE COLOR
Examine the specimen under a good light source, looking down through the container against a White
Background
Urine Clarity
Urine Clarity Refers to the Transparency/Turbidity of a urine specimen
HOW TO EXAMINE URINE CLARITY
 Thoroughly Mix the Specimen
 Examine the specimen while holding in front of a light source
 View through newspaper print

E. Drug Specimen Collection

CHAIN OF CUSTODY (COC), which is a standardized form that must document and accompany
every step of drug testing, from collector to courier to laboratory to medical review officer to employer.
Also, Proper identification of the individual whose information is indicated on the label is required; either
photo identification or positive identification by an employer representative with photo ID is acceptable.
In Specimen collection, it can be labelled “witnessed” or “unwitnessed”. The Required volume of the urine
sample is 30-45ml. Urine temperature should be taken within 4 minutes from the time of collection to
confirm specimen has not been adulterated. The temperature should read within the range of 32.5°C-
37.7°C. In order to prevent adulteration of the specimen, bluing agent is added to the toilet bowl.

IV. Feces/Stool
The laboratory provides patients with several types of containers for collection of fecal (stool)
samples, which the Random samples used for Cultures, & Microscopic examinations of ova & parasites,
cells, blood, fats & fibers are collected. In addition, quantitative testing for fecal fats & urobilinogen, timed
specimens are required and for safety purposes, the outsides of the container should not be contaminated.
V. Semen
In this procedure, Semen samples are collected and tested to evaluate fertility and post vasectomy
procedures. The Patients should abstain for 3 days but no longer than 5 days before collecting the specimen.
The entire Ejaculate must be collected and if the specimen is collected at home, it must be kept warm &
delivered in the lab within 1 hour. In accepting specimens, the phlebotomist must record the time of
collection, & the sample receipt.
Methods of Collection
1. Self-production or Masturbation (Preferred Method)
2. Coitus interruptus (withdrawal method)
3. Vaginal vault aspiration (aspiration of semen from the vaginal vault after coitus)
4. Condom method (use only nonlubricated containing rubber or silastic condom)
VI. Cerebrospinal Fluid
It is routinely collected by lumbar puncture between the third, fourth, or fifth lumbar vertebrae.
Normal CSF appears clear and colorless. Cerebrospinal fluid is collected to diagnose meningitis, subdural
hemorrhage, and other neurological disorders.
CSF Tubes for Examination
TUBE 1 – Chemistry/Serology (least affected by blood or bacteria introduced by spinal tap)
TUBE 2 – Microbiology: Stored at Room Temp
TUBE 3 – Hematology (least likely to contain cells introduced by the spinal tap): Stored at Ref Temp
TUBE 4 (optional) – Micro/Sero (better exclusion of skin contamination)
One Tube is Submitted:
1. Microbiology – to prevent contamination
2. Hematology (Cell Count) – to prevent cell destruction
3. Chemistry/Serology – least likely to be affected by contamination
VII. Serous Fluid
The fluid is located between the parietal membrane and visceral membrane of the pleural,
pericardial, and peritoneal cavities and provides lubrication to prevent the friction between the two
membranes as a result of movement of the enclosed organs.
Samples are collected into EDTA evacuated tubes for cell counts and differential, sterile
heparinized evacuated tubes for microbiology and cytology, and heparinized tubes for chemistry. The type
of fluid should be noted on the sample label.
VIII. Synovial Fluid
Synovial fluid, often referred to as “joint fluid” is a viscous liquid found in the cavities of the
movable or synovial joints that lubricates and reduces friction between bones during joint movement. The
sample is collected via Arthrocentesis
Samples are collected by needle aspiration and collected into tubes based on the required tests: a
sterile heparinized tube for Gram stain and culture and sensitivity, a heparin or EDTA tube for cell counts
and crystal identification, a sodium fluoride tube for glucose analysis, and a non-anticoagulated tube for
other tests.
IX. Amniotic Fluid
The fluid is collected from the fetal sac may be tested for the presence of bilirubin, to monitor
hemolytic disease of the newborn and lipids, to determine fetal lung maturity. Also, it may be examined by
the cytogenetics section for the presence of abnormal chromosomes. Samples for bilirubin analysis must be
protected from light in the same manner as blood samples, and samples for cytogenetic analysis should be
delivered immediately for processing to preserve the limited number of cells present.
X. Sputum
It is a mucus or phlegm collected from the trachea, bronchi, and lungs and is tested for active
tuberculosis (TB) and pneumonia. Sputum is obtained by deep coughing to bring the sputum up from the
lungs and then expelled into a sterile container. The largest amount of sputum is collected in a first morning
sample. Samples should be collected from patients who have abstained from eating, drinking, or smoking.
Before collecting the sample, the patient is asked to rinse his or her mouth with water (do not swallow) to
minimize contamination with saliva. Samples are immediately delivered to the laboratory and kept at room
temperature before processing.

REFERENCE
The Phlebotomy Textbook, Strasinger & Di Lorenzo, 3rd Edition.