Principles of Quality Assurance and Standards

for Veterinary Clinical Chemistry

I. Preanalytical Factors Important in Clinical Chemistry

II. Analytical Factors Important in Clinical Chemistry

• Monitoring
• Method Validation
• Instrumentation
• Quality Control
• Procedures Manual
• Comparison of Test Results

III. Postanalytical Factors Important in Clinical Chemistry

I. Preanalytical Factors Important in Clinical Chemistry [Return to Top]

A. Specimen Collection, Handling, and Transport to the Laboratory

Samples should be appropriately collected, handled and transported to the laboratory in a timely manner, dependent on the type of specimen and its stability. For any assay performed in the laboratory, information concerning sample requirements, proper collection, handling, and delivery or shipping procedures should be available to clients in a laboratory services manual, special information sheets, journal or newsletter articles, other written materials, or by personal or telephone conversation.

B. Specimen Identification

Specimens should be identified with pertinent information as determined by the laboratory (such as owner, species, animal signalment, name of clinic or doctor, address, telephone and fax numbers, e-mail address, location from which the specimen was collected, etc.) on the submission container and submission form.

C. Test Identification

The requested test(s) should be clearly stated on the submission form.

D. Specimen Accessioning

The specimen should be correctly entered into the laboratory system. Test request entry, delivery of the specimen to the correct location, and specimen aliquoting (if necessary) or sharing between laboratories or departments (ie, pharmacology, endocrinology, and clinical chemistry) should be coordinated.

E. Client Communication and Education

Communication between laboratory personnel and clients should be timely and courteous regarding pre-analytical factors influencing laboratory test results (eg, incomplete submission forms, inappropriate sample or sample handling or poor sample quality). Clients should be informed of the expected time for receipt of preliminary and final reports.

F. Personnel Safety

Personal protective equipment should be appropriate for handling specimens and equipment used for clinical chemistry. Safety procedures and disposal of all samples and supplies should be appropriate for the type of specimen. Personnel should receive safety and biohazard training and information about exposure to potentially hazardous chemicals or infectious agents. All training should be documented.

G. Laboratory Environment

The laboratory space should be clean, well lit, and organized to ensure proper achievement of the above goals.

H. Personnel Requirements

Laboratory personnel should have training in specimen handling and sample preparation. Documentation of training, continuing education and periodic proficiency assessment should be at the discretion of the laboratory director.

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II. Analytical Factors Important in Clinical Chemistry [Return to Top]

A. Monitoring

1. Internal monitoring should include the following:

(a) Quality of water (as specified by instrumentation and assays).

(b) Stability of electrical power (as specified by instrumentation).

(c) Temperatures of water bath, refrigerator, and freezer (recommended at least monthly).

(d) Regular calibration of analytical balances and pipettes (recommended annually).

(e) Maintenance of up-to-date procedure manuals with clearly stated dates when procedures are first implemented and when any changes are made and implemented.

(f) Maintenance of adequate inventory, with proper storage and handling.

(g) Maintenance of a log of changes in any procedures, problems or other factors affecting methods, as well as actions that resolved the problem. All entries should be clearly dated and signed by laboratory personnel.

2. External monitoring should include participation in an external proficiency program

(a) All participating laboratories should analyze the same materials.

(b) Results should be tabulated regularly (monthly, quarterly) and distributed to participants with statistical summaries and comparison of participating laboratories with mean indices expressing the closeness of individual laboratory results to the group mean.

(c) Means should be calculated and analyzed based on identification of the method (same methods compared).

(d) Each laboratory should carefully assess the validity of their reported performance and consider any changes indicated by the proficiency program.

B. Method Validation (Return to Top)

Method validation should be performed before a test procedure is placed into routine use. Validation may be accomplished by thoroughly testing reference materials or by comparison of results of tests performed by an alternative method. For each method, the laboratory should verify the manufacturer’s claims and any adjustments before initiating patient testing.

Method validation should provide evidence of the following:

1. Accuracy – Perform either (a) or (b)

(a) Run known value substance and compare results to expected value.

(b) Perform split sample patient comparison between existing method of known accuracy and new method .

2. Precision – Perform either (a) or (b)

(a) Run 10 replicates of 2 levels of quality control (QC) samples.

(b) Gather 21 results; 7 results in each of 3 separate runs (better estimate of day-to-day precision, as well as within-run precision).

With results from (a) or (b) determine mean, standard deviation (SD) and coefficient of variation (CV). Determine whether within-run SD is acceptable.

3. Sensitivity – Perform (a), (b) or (c)

(a) Assess manufacturer’s claims.

(b) Use concentration of low calibrator or another sample or fluid with low levels of analyte.

(c) Run a series of dilutions and assess acceptability of performance.

4. Specificity – Perform (a) or (b)

(a) Use published list of interfering substances, check with manufacturer.

(b) Assess known or suspected interfering substances by spiking specimens or use patient material with known conditions.

5. Linear reportable range

(a) Establish upper and lower limits for reporting patient values based on calibration materials.

(b) For the lower limit, there should be confirmation of the discriminatory ability of the test.

(c) The highest calibration point is the maximum upper limit and the lowest calibration point or zero should be the minimum lower limit for reporting patient results.

6. Linearity – Perform either (a) or (b)

(a) Determine by analyzing multiple dilutions of either a high calibrator, control or patient samples with increased levels of analyte.

(b) Analyze calibrators of variable, known concentrations.

(c) Linearity should be established at the time of validation and whenever new or altered reagents are used.

7. Reference intervals

(a) The laboratory should establish or validate existing reference intervals for each method and species before reporting results.

(b) Parallel tests should be run to confirm reference intervals for controls when changing reagents or QC lot number.

C. Instrumentation (Return to Top)

1. Instrument performance

The equipment and instrument used must be capable of providing test results within the laboratory’s stated performance characteristics. These include:

(a) Detection limits

(b) Precision

(c) Accuracy

(d) Specificity

(e) Sensitivity

(f) Freedom from interferences and related test variables (refer to previous section on method validation)

(g) Additional points to consider:

• Instruments with adjustable setting for different substances and/or species should be carefully checked for compliance
• Compare and make adjustments for performance characteristics as defined by the laboratory and the manufacturer
• Make certain species differences are accommodated

2. Function checks

(a) Appropriate function checks should be made on all instruments. These are critical operating characteristics of an instrument, ie, stray light, zeroing, electrical levels, optical alignment, background checks, etc.

(b) Laboratory personnel should recheck and/or calibrate each instrument daily or once per shift, prior to patient testing, to ensure that it is functioning correctly and is properly calibrated. This includes daily QC.

3. Calibration

(a) Instruments should be calibrated every 6 months or more frequently if indicated by:

• Manufacturer’s recommendation
• After major service
• QC outside limits or troubleshooting indicates need
• Laboratory determination that volume, equipment performance or reagent stability indicate a need for more frequent calibration

(b) After calibration, controls should be run

4. Laboratory personnel knowledge of equipment and its use, including, but not limited to:

(a) Linearity differences from possible manufacturer’s range (human) to animal

(b) Effects of hemolysis, lipemia, icterus, caretenoid pigments (especially large animals), and different anticoagulants on each assay

(c) Reportable ranges

(d) Species-specific ranges and reference intervals

(e) Expected abnormal ranges

(f) Common problems encountered with veterinary samples

(g) Regular instrument maintenance schedule

(h) Replacement of inadequate or faulty equipment

(i) Problem-solving procedures, troubleshooting

D. Quality Control (Return to Top)

1. For each run, at lest 2 controls should be assayed. Use of "high" and "low" abnormal controls is recommended.

2. Maximum length of a run is 24 hours. If the instrument manufacturer requires more frequent controls, observe the recommended frequency (ie, some blood gas instruments).

3. Verify that the instrument is stable over the "run time". During a validation check, controls are assayed more frequently to establish run time.

4. Establish QC frequency; consider the following:

(a) Test volume (number performed each run or day) and frequency

(b) Technique dependence of the method

(c) Analyte or reagent stability

(d) Frequency of QC failures

(e) Training and experience of personnel

(f) Cost of QC (increasing frequency adds to cost-per-test)

5. Quality control parameters

(a) Mean, SD and CV should be calculated (minimum n = 20).

(b) Controls should be assayed in the same manner as patient specimens .

(c) A mechanism should be in place to determine whether testing personnel follow policies and procedures correctly.

(d) Use of Westgard multirule procedures or other rules based on QC validation is recommended.

(e) Policies and procedures should be written and available in a laboratory Standard Operating Procedures (SOP) manual to ensure accurate and reliable test results .

(f) An SOP manual should have clearly marked and dated entries of current procedures (manufacturer package inserts are sufficient as long as verified) and when any changes are made and implemented.

(g) QC records should be reviewed frequently to ensure that when QC values fail to meet the criteria for acceptability, suitable action is taken.

(h) Control products should be purchased commercially, if possible. If using calibrators as controls, use a different lot as QC material. If patient pooled samples are used, establish the mean value of all analytes (minimum n = 10 to establish a mean).

(i) Monitor results of clinical specimens for various sources of error by use of parameters such as anion gap, comparison of test results with previous submissions from same patient (delta checks), and investigation of markedly abnormal results (limit checks).

E. Procedures Manual (Return to Top)

1. All procedures currently in use should be included. Protocols may be organized in manuals and/or stored in computers, and be written form. They should contain such information as:

(a) Patient preparation

(b) Specimen collection, processing and handling

(c) Criteria for rejection of specimens

(d) Limitations of and things that interfere with the method in use

(e) Step-by-step procedures

(f) Reagent preparation or manufacturer

(g) Reference interval

(h) Reportable range

(i) Literature references

(j) Reagent labeling: content, storage requirements, expiration

(k) Laboratory-specific information, such as:

• Identification of instrument used
• Result reporting method
• Actions to take when system is down
• Criteria for specimen referrals to outside laboratories ("send outs")
• Quality control procedures
• Documentation of critical values
• Clearly stated and dated entries of procedure implementation or change

F. Comparison of Test Results (Return to Top)

If the laboratory performs the same test by more than 1 method or at more than 1 test site, or the test is sometimes also sent to a referral laboratory, comparisons should be run at least twice annually to define the relationships between methods and sites. Comparison of different test methods for the same analyte within the laboratory or between laboratories (if samples are tested in-house and at a referral laboratory) is recommended. This should be done every 6 months or at a frequency determined by the laboratory manager. The following steps should be included:

1. Perform a 20-sample or greater comparison using specimens covering the analytical range.

(a) Group data in an x-y comparison plot

(b) Calculate slope and intercept by a least squares method

2. Laboratory director or qualified personnel should define acceptable performance limits .

3. If individual test results performed on the same patient or material do not correlate with each other (ie, BUN/creatinine, electrolyte balance), the cause should be investigated and corrective action taken.

Postanalytical Factors Important in Clinical Chemistry [Return to Top]

A. Computer Entry of Data

Reports should be accurate whether created manually or electronically, and in a standard format as established by the laboratory. Established laboratory standards for uniform reporting should be met.

B. Report Generation

Reports should be in a format that is readable and easily understood, with appropriate references or explanations as needed. They should be generated in a timely manner relative to preanalytical and analytical components.

C. Report Delivery

Report delivery should be timely, to the correct client, and in a manner agreed upon by the client and the laboratory.

D. Specimen Storage

Specimens should be stored under appropriate conditions for a predetermined time period, as determined by specimen stability, laboratory policy and/or certification/accreditation requirements.

E. Specimen Disposal

Laboratories should appropriately dispose of biohazardous and non-biohazardous materials and specimens, including timely emptying of all containers and trash bins.

F. Personnel Safety

Conditions should be appropriate for computer entry, transcription, handling of specimens, specimen disposal and all other postanalytical tasks.

G. Laboratory Environment

Laboratory environment should meet standard requirements necessary for safe, rapid, efficient and effective performance.

H. Personnel Requirements

Personnel should meet training requirements as indicated for specific areas of the laboratory.

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