• All samples must be considered to be infectious

Sample preparations for laboratory investigations

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Biological samples may contain infective agents and must be handled with care

• All samples must be considered to be infectious

• Use of “Universal Precautions” handling

• Never assume any sample is “safe”

• Today’s symptoms may be tomorrow’s diagnosis of

infection

Biological Specimens

Any biological material taken from a patient for diagnostic, prognostic or therapeutic monitoring

• Blood

• Urine

• Saliva

• sweat

• Semen

• Amniotic Fluid

• Synovial Fluid

• Cerebrospinal Fluid

• Gastric Juice

• Stools

• Gall stone

• Kidney Stone

• Tissue Specimen

Preparation of Blood Sample

One of three different specimens may be used:

• whole blood

• serum

• Plasma

Whole-blood samples:

It must be analyzed within limited time (why?)

– Over time, cells will lyse in whole-blood which will change the conc. of some analytes as potassium, phosphate and lactate dehydrogenase.

– Some cellular metabolic processes will continuo which will

alter analytes conc. like glucose and lactate.

Serum

Difference between Serum and plasma:

• Serum is the same as plasma except it

doesn't contain clotting factors (as fibrin).

• Plasma contains all clotting factors.

• So, serum and plasma all has the same

contents of electrolytes, enzymes proteins, hormones except clotting factors

• Serum is mainly use in chemistry lab &

serology.

Procedure of Serum preparation

• Draw blood from patient. Select vacutainer with no anticoagulant.

• Allow to stand for 20-30 min for clot formation.

• Centrifuge the sample to speed separation and affect a greater packing of cells. Clot and cells will separate from clean serum and settle to the

bottom of the vessel.

• The supernatant is the serum which can be now collected by

• Dropper or pipette for testing purposes or stored (-

20°C to -80°C) for subsequent analysis or use.

Plasma

• The tube will have anti-coagulation

• After centrifugation the blood sample got

separated into three layers

Procedure of plasma preparation

• Draw blood from patient. Select the tube with an appropriate anticoagulant.

• Mix well the blood with anticoagulant.

• Allow to stand for 10 min.

• Centrifuge the sample to speed separation and affect a greater packing of cells.

• The supernatant is the plasma which can be now collected for testing

• Purposes or stored (-20°C to -80°C) for subsequent analysis or use.

Blood Collection

Collection Tubes

• The most widely used tubes for blood

collection are evacuated tubes (Vacutainers) – Negative pressure facilitates collection

– Easy to use – Sterile

– Universally used colour-coded rubber stoppers to denote tube type.

– Tubes can contain various anticoagulants for the collection of whole blood or

plasma.

– Tubes can have additives for specific tests

(glucose, metals)

Vacutainer blood collection

Venipuncture

Anticoagulants

• Sodium Citrate

• Potassium Oxalate

• Heparin

• EDTA (Ethylene Diamine Tetraacetic Acid)

• Acid Citrate Dextrose (ACD)

Top Color Additives Principle Uses

Lavender EDTA -The strongest anti-coagulant - Ca⁺² chelating agent

- To preserve blood cells components

- Hematology

- Blood bank (ABO) - HbA1C

(Glycosylated Hb)

Light Blue Sodium Citrate Ca⁺² chelating agent - PT: Prothrombin Time

- PTT: Partial

Thromboplastin Time ( in case of

unexplained bleeding and liver disease)

Green Sodium

Heparin or Lithium Heparin

Heparin binds to Thrombin and inhibits the second step in the coagulation cascade

(Prothrombin Thrombin)

Fibrinogen Fibrin

Enzymes Hormones

Electrolytes (Na⁺, K⁺, Mg⁺, Cl^-

Heparin

Plasma Separating Tubes (PST)

Top Color Additives Principle Uses Black Sodium Citrate Ca⁺² chelating

agent

ESR ( Erythrocyte Sedimentation Rate)

to test how much inflammation in the patient, unexplained fever, Arthritis, Autoimmune Disorder Gray -Sodium Fluoride

-Potassium Oxalate

Glycolysis inhibitor

Anti-Coagulant

Glucose tests

Royal Blue Heparin Na-EDTA

Anti-Coagulant Tube should not be

contaminated with metals

Toxicology

Trace Elements and metals

Yellow ACD ( Acid-Citrate Dextrose)

Anti-Coagulant DNA Studies Paternity Test HLA Tissue Typing

(Human Leukocyte Antigen) The body used this protein to

differentiate the self-cells from non- self cells

Equipments

ESR H.V.

Centrifuge

Urine analysis

•

18

Urine collection

• 24 hour sample must include all urine passed in this period

• If less than 24h, inform the lab

• Mid stream sample

• Early morning sample – often best

• Correct container type

− Macroscopic Examination

− Chemical Analysis (Urine Dipstick)

− Microscopic Examination

− Culture (not covered in this lecture)

− Cytological Examination

Urine examination

Stool Analysis

Stool

5. Epithelial cells that have been shade

1. Waste residue of indigestible material (cellulose during the previous 4 days)

2. Bile pigments and salts

3. Intestinal secretions, including mucus

4. Leukocytes that migrate from the bloodstream

6. Bacteria and Inorganic material(10-20%) chiefly calcium and phosphates.

Undigested and unabsorbed food .

Random Collection

1. Universal precaution

2. Collect stool in a dry clean container

3. uncontaminated with urine or other body secretions, such as menstrual blood

4. Collect the stool with a clean tongue blade or similar object.

5. Deliver immediately after collection

Ova and parasites collection

1. Warm stools are best for detecting ova or parasites.

Do not refrigerate specimen for ova or parasites.

2. If the stool should be collect in 10 % formalin or PVA fixative, storage temperature is not critical.

3. Because of the cyclic life cycle of parasites, three

separate random stool specimens are recommended.

Enteric pathogen collection

1. Some coliform bacilli produce antibiotic substances that destroy enteric pathogen.Refrigerate specimen immediately.

2. A diarrheal stool will usually give accurate results.

3. A freshly passed stool is the specimen of choice.

4. Stool specimen should be collected before antibiotic therapy, or as early in the course of the disease.

5. If blood or mucous is present, it should be included in the specimen