SEROLOGY

MODULE 9
 SEROLOGY: It is the study of antigen and antibody reaction

When to use serology?

 Unable to culture infectious agent
 Confirmation of etiologic agent
 Diagnosis of immunologically-related
disorders
 Determine immune status
 INTRODUCTION TO SEROLOGY

• SAFETY IN IMMUNOLOGY • QUALITY ASSURANCE • BASIC SEROLOGIC

SEROLOGY LABORATORY AND CONTROL LABORATORY
TECHNIQUES
 Safety in Immunology
Serology Laboratory
 SAFETY STANDARDS AND AGENCIES • Department of Health
• Occupational Safety and Health through the Health
Administration (OSHA) Facilities and Services
• Clinical and Laboratory Standards Regulatory Bureau.
Institute (CLSI) • Department of Labor
• Centers for Disease Control and and Employment
Prevention (CDC) (DOLE)
• College of American Pathologists
(CAP)
• Joint Commission on Accreditation
of Healthcare Organizations (JCAHO)
 Safety in Immunology
Serology Laboratory
 UNIVERSAL PRECAUTIONS
• Under universal precautions, all patients are assumed to be
possible carriers of bloodborne pathogens
• instituted by the CDC in 1985 to protect health-care workers
from exposure to bloodborne pathogens, primarily hepatitis
B virus (HBV) and HIV
 Safety in Immunology
Serology Laboratory
 BODY SUBSTANCE ISOLATION (BSI)
• It is a medication of universal precaution where it is not limited to
bloodborne pathogens and considers all body fluids and moist
body substances to be potentially infectious.
• Personnel should wear gloves at all times when encountering moist
body substances. A disadvantage of the BSI guideline is that it does
not recommend handwashing after removing gloves unless visual
contamination is present.
• The major features of UP and BSI have now been combined and are
called standard precautions.
 Safety in Immunology
Serology Laboratory
 STANDARD PRECAUTION
• all human blood and other body fluids are treated as
potentially infectious for HIV, Hepatitis B virus and other
blood-borne microorganism.
**BLOOD-is the most important source of HIV, HBV and
other blood borne pathogens in the occupational settings.
 Safety in Immunology
Serology Laboratory
 STANDARD PRECAUTION
• Compliance with the OSHA Bloodborne Pathogens Standard and
the Occupational Exposure Standard is required to provide a
safe work environment. OSHA mandates that the employer do
the following:
• Educate and train all health care workers in Standard Precautions
and in preventing bloodborne infections.
• Provide proper equipment and supplies (e.g., gloves).
• Monitor compliance with protective biosafety policies.
 Safety in Immunology
Serology Laboratory
 THINGS TO REMEMBER:
 HBV can be present in extraordinarily high
concentrations in blood, but HIV is usually found in
lower concentrations.
 HBV may be stable in dried blood and blood products at
25°C for up to 7 days.
 HIV retains infectivity for more than 3 days in dried
specimens at room temperature (20-24C) and for
more than 1 week in an aqueous/wet environment at
room temperature.
 Safety in Immunology
Serology Laboratory
 THINGS TO REMEMBER:
 Both HBV and HIV may be transmitted indirectly.
 Viral transmission through contact to inanimate
objects
 Virus is transmitted through skin and mucous
membrane by hand contact contaminated surface and
non intact skin or mucous membrane
 The mode of transmission of HBV and HIV is the same
although the potential for transmission in
occupational setting is greater for HBV than HIV.
 Safety in Immunology
Serology Laboratory
 THINGS TO REMEMBER:
 The modes of transmission for HBV and HIV are similar, but
the potential for transmission in the occupational setting is
greater for HBV than HIV.
 Both HBV and HIV may also be transmitted directly.
 Percutaneous (parenteral) inoculation of blood,
plasma, serum or certain body fluids from accidental
needlestick
 Contamination of skin with blood or certain body fluids
without overt puncture, caused by scratches,
abrasions, burns, weeping or exudative skin lesions.
 Safety in Immunology
Serology Laboratory
 THINGS TO REMEMBER:
 Both HBV and HIV may also be transmitted directly.
 Exposure of mucous membranes (oral, nasal or
conjunctival) to blood or certain body fluids as the
direct result of pipetting by mouth, splashes, or
spattering.
 Centrifuge accidents or the improper removal of
rubber stoppers from test tubes, producing droplets.
If these aerosol products are infectious and come into
direct contact with mucous membranes or nonintact
skin, direct transmission of virus can result.
 Safety in Immunology
Serology Laboratory
 THINGS TO REMEMBER:
 The likelihood of infection in health care workers after
exposure to blood infected with HBV or HIV depends on the
following factors:
 Concentration of HBV or HIV (viral concentration is higher
for HBV than HIV)
 Duration of the contact
 Presence or skin lesions or abrasions on the hands or
exposed skin of the health care worker
 Immune status of the health care worker for HBV
 Safety in Immunology
Serology Laboratory
 THINGS TO REMEMBER:
 Studies have demonstrated that HIV is inactivated
rapidly after being exposed to common germicides at
concentrations much lower than used in practice.
 Diluted household bleach (10% hypochlorite)
prepared daily inactivates HBV in 10 minutes and
HIV in 2 minutes.
 Disposable materials contaminated with blood
should placed in containers marked with
“BIOHAZARD” and properly discarded.
 Quality Assurance and
Control
CLINICAL LAB REGULATING AND ACCREDITING AGENCIES
 Clinical Laboratory Improvement Amendments of 1988 (CLIA
’88)
 established a minimum threshold for all aspects of clinical
laboratory testing
 The Joint Commission (TJC),
 Commission on Office Laboratory Accreditation
 College of American Pathologists
 International Organization for Standardization (ISO)
 ISO 15189
 Quality Assurance and
Control
 Non-analytical factors related to testing Accuracy
 Qualified Personnel
 Established Laboratory Policies
 Laboratory Procedure Manual
 Test Requisitioning
 Patient Identification, Specimen Procurement, and
Labeling
 Written
Procedure
Protocol
 Quality Assurance and
Control
 Non-analytical factors related to testing
Accuracy
• Preventive Maintenance of Equipment
• Appropriate Testing Methods
• Inaccurate Results
 Quality Assurance and
Control
 Errors related to Phase of testing
 Most laboratory errors are related to the preanalytical and
postanalytical phases of testing
 Possible
Causes of
Technical
Errors
 Quality Assurance and
Control
 Important Definitions
 Quality control-consists of procedures used to detect errors that
result from test system failure, adverse environmental
conditions, and differences between technologists, as well as the
monitoring of the accuracy and precision of test performance
over time.
 Accuracy describes how close a test result is to the true value.
 Precision describes how close the test results are to one another
when repeated analyses of the same specimen are performed
 Quality Assurance and
Control
 Important Definitions
 Sensitivity of a test-defined as the proportion of subjects with the specific
disease or condition who have a positive test result.
 Quality Assurance and
Control
 Important Definitions
 Specificity of a test-defined as the proportion of subjects without the
specific disease or condition who have a negative test result (i.e., assay
correctly excludes with a negative result)
 Quality Assurance and
Control
 Proficiency Testing
 A form of external assessment of laboratories to ensure
their ability to perform to the level of competence and
quality required.
 A laboratory tests a specimen is provided by a government
agency, professional society, or commercial company.
These identical samples are sent to a group of laboratories
participating in the PT program. Each laboratory analyzes
the specimen, reports the results to the agency, and is
evaluated and graded on those results compared with
results from other laboratories. In this way, quality control
between laboratories is monitored
 Quality Assurance and
Control
 Control Specimens
 specimen with a known value that is similar in
composition to the patient’s blood
 A control specimen must be carried through the entire
test procedure and treated in exactly the same way as
any unknown specimen
 Quality Assurance and
Control
 Control Specimens
 If the control value in a determination is out of the acceptable range (out of
control), one or more of the following factors may be responsible:
1. Deterioration of reagents or standards
2. Faulty instrument or equipment
3. Dirty glassware
4. Lack of attention to timing or incubation temperature
5. Use of a method not suited to the needs and facilities of the laboratory
6. Use of poor technique by the technologist doing the test
7. Statistics: a certain percentage of all determinations will be statistically out
of control
 Quality Assurance and
Control
 Reference Range
 In analytical immunology and serology testing using
methods such as enzyme immunoassay, quantitative
reference range statistics can be used
 the range of values that includes 95% of the test results
for a healthy reference population
 Replaces normal values, or normal range
 Basic Serological
Laboratory Technique
 Specimen Preparation and Processing
 The most frequently encountered specimen in immunological
testing is serum
 Care must be taken to avoid hemolysis, since this may produce
false positive result.
 The blood specimen is allowed to clot at room temperature or at
4C and then centrifuged
 Serum should be promptly separated into another
tube without transferring any cellular elements.
 Fresh, non heat inactivated serum is usually recommended for
testing
 Basic Serological
Laboratory Technique
 Specimen Preparation and Processing
 If testing cannot be performed immediately, serum may be stored
between 2C and 8C for up to 72 hours.
 If there is any additional delay in testing, the serum should be
frozen at -20C or below
 In obtaining specimens for serologic testing, it is important to
consider the phase of the disease and the condition of the patient
at the time of specimen collection. This is especially important in
assays for diagnosis of infectious diseases
 Acute serum-sample drawn during the acute phase of illness when
the disease is first discovered or suspected
 Convalescent serum-sample drawn during convalescent phase,
usually about 2 weeks later
 Basic Serological
Laboratory Technique
 Specimen Preparation and Processing
 A differencein the amount of antibody
present, or the antibody titer, may be
noted when two different samples are
tested concurrently.
 Some infections, such as Leigionnaires disease
or hepatitis, may not manifest a rise in titer in
months after the acute infection.
 Basic Serological
Laboratory Technique
 Specimen Preparation and Processing
 Most immunology tests are done on serum, although body fluids may
also be tested.
 Lipemia, hemolysis, or any bacterial contamination can make the
specimen unacceptable.
 Icteric or turbid serum may yield valid results for some tests but may
interfere with others. Blood specimens should be collected before a
meal to avoid the presence of chyle, an emulsion of fat globules that
often appears in serum after eating, during digestion.
 Contamination with alkali or acid must be avoided because these
substances have a denaturing effect on serum proteins and make the
specimens useless for serologic testing.
 Basic Serological
Laboratory Technique
 Pipettes
 Pipettes are used in the immunology-serology
laboratory for the quantitative transfer of
reagents and the preparations of serial dilutions
of specimens such as serum
 Basic Serological
Laboratory Technique
 Graduated Pipettes
 delivers a particular amount of liquid
 Has several graduation, or calibration, marks
 used primarily for measuring reagents but are not calibrated with
sufficient tolerance for measuring standard or control solutions,
unknown specimens, or filtrates.
 A graduated pipette is a straight piece of glass tubing with a tapered
end and graduation marks on the stem separating it into parts
 These pipettes come in various sizes, or capacities,including 0.1, 0.2,
1.0, 2.0, 5.0, 10, and 25 mL
 used when speed is more important than precision
 Basic Serological
Laboratory Technique
 Serologic Pipettes
 recognized by a frosted ring at the noncalibrated
end, with calibrations extending to the tip
 The letters TD (to deliver) appear on the pipette
and, for quick recognition, each size of pipette has
an imprinted, color-coded band that indicates the
volume
 allowed to empty by gravity, depending on the
calibration, the remaining drop needs to be
expelled to deliver the full volume
 Basic Serological
Laboratory Technique
 Automatic Pipettes
 Automatic pipettes allow fast, repetitive
measurement and delivery of solutions of equal
volumes.
 mechanically operated and uses a piston-
operated plunger
 These are adjustable so that varying amounts of
reagent or sample can be delivered with the
same device.