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Principles of basic techniques and laboratory safety
Abstract
Background
To appropriately interpret clinical laboratory test results and adequately validate assays, the basic principles and techniques of analytical chemistry need to be understood. These techniques should be used by laboratory professionals in a safe testing environment.
Content
Factors that affect the analytical process and operation of the clinical laboratory are described in this chapter. The concepts of solute and solution, and the international system of units used to standardize their expression and reporting are described. These solutions are composed of various types of chemicals used in the development of clinical laboratory assays. The importance of water purity, appropriate reagent preparation, and the different types of reference materials are addressed. The principles of basic techniques in the clinical laboratory, including pipetting, centrifugation, radioactivity, gravimetry, and thermometry, are also discussed. These techniques are used in a variety of laboratory tasks such as making buffers, performing dilutions, evaporation, lyophilization, and filtration. Safety is a constant and crucial concern for laboratory personnel. Each laboratory must create a comprehensive safety program. Plans for the handling of chemicals, exposure to blood-borne pathogens, tuberculosis, and other highly infectious agents are necessary components of a safety plan. The training of laboratory personnel to identify various types of biological, chemical, and electrical hazards and to react appropriately to fire must be addressed in such a plan.
Concept of solute and solvent
Many analyses in the clinical laboratory are concerned with determination of the presence or measurement of the concentration of substances in solutions; these solutions are most often blood, serum, urine, spinal fluid, or other bodily fluids (see Chapter 4 ).
A solution is a homogeneous mixture of one or more solutes dispersed molecularly in a sufficient quantity of a dissolving solvent. In laboratory practice, solutes are typically measured and are frequently referred to as analytes or measurands. A solution may be gaseous, liquid, or solid. A clinical laboratorian is concerned primarily with the measurement of gases or solids in liquids, in which there is always a relatively large amount of solvent in comparison with the amount of solute.
Expressing concentrations of solutions
The following equations define the expression of concentrations:
In the past, milliequivalent (mEq) was used to express the concentration of electrolytes in plasma. Currently, the recommended unit for expressing the concentration of an electrolyte in plasma is millimoles per liter (mmol/L). For example, if a sample contains 322 mg of sodium per deciliter, then the serum contains 3220 mg/L. The molar concentration of Na is
In clinical laboratory practice, a titer is the lowest dilution at which a particular reaction takes place. Titer is customarily expressed as a ratio (e.g., 1:10 or 1 to 10).
Regarding gases in solution, Henry’s law states that the solubility of a gas in a liquid is directly proportional to the pressure of the gas above the liquid at equilibrium. Thus as the pressure of a gas is doubled, its solubility is also doubled. The relationship between pressure and solubility varies with the nature of the gas. When several gases are dissolved at the same time in a single solvent, the solubility of each gas is proportional to its partial pressure in the mixture. The solubility of most gases in liquids decreases with an increase in temperature, and boiling a liquid frequently drives out all dissolved gases. Using the Système International d’Unités (SI), gas concentrations are expressed in moles per cubic meter (mol/m 3 ).
Units of measurement
A meaningful measurement is expressed with both a number and a unit. The unit identifies the dimension—mass, volume, or concentration—of a measured property. The number indicates how many units are contained in the property.
Traditionally, measurements in the clinical laboratory have been made in metric units. In the early development of the metric system, units were referenced to length, mass, and time. The first absolute systems were based on the centimeter, gram, and second, and then the meter, kilogram, and second. The SI is a different system that was accepted internationally in 1960. The units of the system are called SI units.
International system of units
Base, derived, and supplemental units are the three classes of SI units. The eight fundamental base units are listed in Table 15.1 . A derived unit is derived mathematically from two or more base units ( Table 15.2 ). A supplemental unit is a unit that conforms to the SI but has not been classified as either base or derived. At present, only the radian (for plane angles) and the steradian (for solid angles) are classified this way.
TABLE 15.1
Standard International Base Units
| Quantity | Name | Symbol |
|---|---|---|
| Length | Meter | m |
| Mass | Kilogram | kg |
| Time | Second | s |
| Electrical current | Ampere | Å |
| Thermodynamic temperature | Kelvin | K |
| Amount of substance | Mole | mol |
| Luminous intensity | Candela | Cd |
| Catalytic amount | Katal | kat |
TABLE 15.2
Examples of Standard International-Derived Units Important in Clinical Medicine, Expressed in Terms of Base Units
| Quantity | Name | SI Symbol | Expression in Terms of SI Base Units | Expression in Terms of Other SI Units |
|---|---|---|---|---|
| Volume | Cubic meter | m 3 | m 3 | |
| Mass density | Kilogram per cubic meter | kg/m 3 | kg/m 3 | |
| Concentration of amount of substance | Mole per cubic meter | mol/m 3 | mol/m 3 | |
| Frequency | Hertz | Hz | s −1 | |
| Force | Newton | N | m · kg · s −2 | |
| Pressure | Pascal | Pa | m −1 · kg · s −2 | N/m 2 |
| Energy, work, quantity of heat | Joule | J | m 2 · kg · s −2 | N · m |
| Power | Watt | W | m 2 · kg · s −3 | J/s |
| Electric potential, potential difference, electromotive force | Volt | V | m 2 · kg · s −3 · Å −1 | W · Å −1 |
SI, International System of Units.
The Conférence Générale des Poids et Mésures recognizes that some units outside the SI unit continue to be important and useful in particular applications. An example is the liter as the reference volume in clinical analyses. Liter is the name of the submultiple (cubic decimeter) of the SI unit of volume, the cubic meter. Considering that 1 m 3 represents approximately 200 times the blood volume of an adult human, the SI unit of volume is neither a convenient nor a reasonable reference volume in a clinical context. Nevertheless, the Conférence Générale des Poids et Mésures recommends that such exceptional units as the liter should not be combined with SI units and preferably should be replaced with SI units whenever possible.
The minute, hour, and day have had such long-standing use in everyday life that it is unlikely that new SI units derived from the second could supplant them. Some other non-SI units are still accepted, although they are rarely used by most individuals in their daily lives, but they have been important in some specialized fields. Details of the SI system can be found in an expanded version of this chapter.
In practical application of units, certain values are too large or too small to be expressed conveniently. Numeric values are brought to convenient size when the unit is appropriately modified by official prefixes ( Table 15.3 ).
TABLE 15.3
Metric Prefixes of Standard International Units a
From The International System of Units (SI). Washington, DC: National Institute of Standards and Technology, 1991.
| Factor | Prefix | Symbol |
|---|---|---|
| 10 24 | Yotta | Y |
| 10 21 | Zetta | Z |
| 10 18 | Exa | E |
| 10 15 | Peta | P |
| 10 12 | Tera | T |
| 10 9 | Giga | G |
| 10 6 | Mega | M |
| 10 3 | Kilo | K |
| 10 2 | Hector | H |
| 10 1 | Deka b | Da |
| 10 −1 | Deci | D |
| 10 −2 | Centi | C |
| 10 −3 | Milli | m |
| 10 −6 | Micro | μ |
| 10 −9 | Nano | N |
| 10 −12 | Pico | p |
| 10 −15 | Femto | F |
| 10 −18 | Atto | A |
| 10 −21 | Zepto | Z |
| 10 −24 | Yocto | Y |
a The Eleventh Conférence Générale des Poids et Mésures (CGPM) (1960, Resolution 12) adopted a first series of prefixes and symbols of prefixes to form the names and symbols of the decimal multiples and submultiples of standard international (SI) units. Prefixes for 10 −15 and 10 −18 were added by the 12th CGPM (1964, Resolution 8), those for 10 15 and 10 18 were added by the 15th CGPM (1975, Resolution 10), and those for 10 21 , 10 24 , and 10 −24 were proposed by the Comité International des Poids et Mésures (CIPM) (1990) for approval by the 19th CGPM (1991).
b The spelling “deca” is used extensively outside the United States.
Standardized reporting of test results
To describe test results properly, it is important that all necessary information be included in the test description. Systems developed for expressing results produced by the clinical laboratory include the Laboratory Logical Observation Identifiers, Names, and Codes (Lab LOINC), and the Nomenclature, Properties, and Units (NPU) developed by the International Federation of Clinical Chemistry and Laboratory Medicine/International Union of Pure and Applied Chemistry (IFCC/IUPAC).
Laboratory logical observation identifiers, names, and codes system
The Lab LOINC system is a universal coding system for reporting laboratory and other clinical observations to facilitate electronic transmission of laboratory data within and between institutions ( http://www.loinc.org ). It has several thousand observations in its database. For each observation, there is a code, a long formal name, a short 30-character name, and synonyms. A mapping program termed “Regenstrief LOINC Mapping Assistant” (RELMA) is available to map local test codes to LOINC codes and to facilitate searching of the Lab LOINC database. Both Lab LOINC and RELMA are available at no cost from http://loinc.org/relma (accessed February 19, 2020).
Nomenclature, properties, and units
The NPU system recommends that the following items be included with each test result:
- 1.
The name of the system or its abbreviation
- 2.
A dash (two hyphens)
- 3.
The name of the analyte (never abbreviated) with an initial capital letter
- 4.
A comma
- 5.
The quantity name or its abbreviation
- 6.
An equal sign
- 7.
The numeric value and the unit or its abbreviation
Applications
The Lab LOINC and NPU coding systems are used in context with existing standards, such as the Systematized Nomenclature of Medicine, Clinical Terms (SNOMED CT). Other such coding systems are the ASTM E1238 (American Society for Testing and Materials), HL7 version 2.2 (Health Level Seven; http://www.hl7.org ; accessed February 19, 2020), and CEN ENV 1613—a standard developed by the European Committee for Standardization of the Comité Européen de Normalisation (CEN) Technical Committee 251 ( http://www.cen.eu/ accessed February 19, 2020) .
SNOMED CT is a comprehensive clinical terminology, originally created by the College of American Pathologists (CAP) and, as of April 2007, owned, maintained, and distributed by the International Health Terminology Standards Development Organisation (IHTSDO) in Denmark ( http://www.ihtsdo.org ; accessed February 19, 2020). IHTSDO is a not-for-profit association that develops and promotes use of SNOMED CT to support safe and effective health information exchange. In practice, the CAP continues to support SNOMED CT operations under contract to the IHTSDO and provides SNOMED-related products and services as a licensee of the terminology.
On April 1, 2009, the owners of LOINC, NPU, and SNOMED CT began a 6-month operational trial of prospective divisions of labor in the generation of laboratory test terminology content. This trial provided practical experience and important information on opportunities to decrease duplication of effort in the development of laboratory test terminology and to ensure that SNOMED CT works effectively in combination with LOINC or NPU. The development of SNOMED CT continues and is critical to the successful implementation of the electronic health record.
Chemicals
The quality of the analytical results produced by the laboratory is a direct indication of the purity of the chemicals used as analytical reagents. The availability and quality of the reference materials used to calibrate assays and to monitor their analytical performance are also important.
Laboratory chemicals are available in a variety of grades. The solutes and solvents used in analytical work are reagent-grade chemicals, among which water is a solvent of primary importance.
Reagent-grade water
Preparation of many reagents and solutions used in the clinical laboratory requires “pure” water. Single-distilled water fails to meet the specifications for Clinical Laboratory Reagent Water (CLRW) established by the Clinical and Laboratory Standards Institute (CLSI). Because the terms deionized water and distilled water describe preparation techniques, they should be replaced by reagent-grade water, and if appropriate, followed by the designation of CLRW, which better defines the specifications of the water and is independent of the method of preparation ( Table 15.4 ).
TABLE 15.4
Clinical and Laboratory Standards Institute Specifications for Clinical Laboratory Reagent Water
From Clinical and Laboratory Standards Institute (CLSI). Preparation and testing of reagent water in the clinical laboratory, 4th ed. CLSI Document GP40-A4-AMD. Wayne, PA: CLSI, 2012.
| CLRW | |
|---|---|
| Microbiological content a (cfu/mL) (maximum) | <10 |
| Resistivity b (MΩ/cm), 25 °C | ≥10 |
| Particulate matter c | Water passed through 0.22-μm filter |
| Total organic content (ng/g) | <500 |
CLRW, Clinical Laboratory Reagent Water.
a Microbiological content. The microbiological content of viable organisms, as determined by total colony count after incubation at 36 ± 1 °C for 14 hours, followed by 48 hours at 25 ± 1 °C, and reported as colony-forming units per milliliters (cfu/mL).
b Specific resistance or resistivity. The electrical resistance in ohms measured between opposite faces of a 1-cm cube of an aqueous solution at a specified temperature. For these specifications, the resistivity will be corrected for 25 °C and reported in megaohms per centimeters (MΩ/cm). The higher the quantity of ionizable materials, the lower will be the resistivity and the higher the conductivity.
c Particulate matter. When water is passed through a membrane filter with a mean pore size of 0.22 μm, nearly all microorganisms and particulates are removed.
Preparation of reagent-grade water
Distillation, ion exchange, reverse osmosis, and ultraviolet (UV) oxidation are processes used to prepare reagent-grade water. In practice, water is filtered before any of these processes are used.
Distillation.
Distillation is the process of vaporizing and condensing a liquid to purify or concentrate a substance or to separate a volatile substance from less volatile substances. It is the oldest method of water purification. Problems with distillation for preparing reagent water include the carryover of volatile impurities and entrapped water droplets that may contain impurities into the purified water. This results in contamination of the distillate with volatiles, sodium, potassium, manganese, carbonates, and sulfates. As a result, water treated by distillation alone does not meet the specific conductivity requirement of CLRW.
Ion exchange.
Ion exchange is a process that removes ions to produce mineral-free deionized water . Such water is most conveniently prepared using commercial equipment, which ranges in size from small, disposable cartridges to large, resin-containing tanks. Deionization is accomplished by passing feed water through columns containing insoluble resin polymers that exchange hydrogen (H + ) and oxygen hydrogen (OH − ) ions for the impurities present in the ionized form in the water. The columns may contain cation exchangers, anion exchangers, or a mixed-bed resin exchanger, which is a mixture of cation- and anion-exchange resins in the same container.
A single-bed deionizer generally is capable of producing water that has a specific resistance in excess of 1 MΩ/cm. When connected in series, mixed-bed deionizers usually produce water with a specific resistance that exceeds 10 MΩ/cm.
Reverse osmosis.
Reverse osmosis is a process by which water is forced through a semipermeable membrane that acts as a molecular filter. The membrane removes 95% to 99% of organic compounds, bacteria, other particulate matter, and 90% to 97% of all ionized and dissolved minerals, but has fewer of the gaseous impurities. Although this process is inadequate for producing reagent-grade water for the laboratory, it may be used as a preliminary purification method.
Ultraviolet oxidation.
UV oxidation is another method that works well as part of a total system. The use of UV radiation at the biocidal wavelength of 254 nm eliminates many bacteria and cleaves many ionizing organics that are then removed by deionization.
Quality, use, and storage of purified water
Autoclave or wash water (formerly type III water) may be used for glassware washing. (However, final rinsing should be done with the water grade suitable for the intended glassware use.) It may also be used for certain qualitative procedures, such as those used in general urinalysis. A variety of purification processes can be used to produce this type of water.
The CLRW, formerly described as type I or II water, is used for most routine clinical laboratory testing. Purification processes for CLRW must achieve specific targets for microbial content, resistivity, particulate matter, and total organic content (see Table 15.4 ). Storage and delivery systems should be constructed to ensure a minimum of chemical or bacterial contamination. The frequency of monitoring of water specifications and the purification system is to be determined by each laboratory.
Special reagent water is used for specific applications. At a minimum, the water should meet CLRW specifications; however, additional parameters may be needed for certain applications. For example, water used for DNA and RNA testing may have specifications for protease and nuclease activity. Water used in trace metal analysis should contain minimally detectable amounts of each metal being measured.
POINTS TO REMEMBER
Water
-
There are several types of water used in the clinical laboratory, including Clinical Laboratory Reagent Water (CLRW), special reagent water, and autoclave and wash water. CLRW has specifications for its microbiological content, resistivity, particulate matter, and total organic content
-
The monitoring of water depends on the type of water and its intended use
Testing for water purity
At a minimum, water should be tested for microbiological content, particulates resistivity, and total organic content. The frequency of monitoring water purity should be established by the laboratory, such that water purity is maintained for its intended purpose. It should be noted that measurements taken at the time of production may differ from those taken at the time and place of use. For example, if the water is piped a long distance, consideration must be given to deterioration en route to the site of use. To meet the specifications for high-performance liquid chromatography (HPLC), in some instances it may be necessary to add a final 0.1-μm membrane filter. The water can be tested by HPLC using a gradient program and monitored with a UV detector. No peaks exceeding the analytical noise of the system should be found.
Reagent-grade or analytical reagent-grade chemicals
Chemicals that meet specifications of the American Chemical Society (ACS) are described as reagent or analytical reagent grade. These specifications have also become the de facto standards for chemicals used in many high-purity applications. These are available in two forms: (1) lot-analyzed reagents, in which each individual lot is analyzed and the actual amount of impurity reported; and (2) maximum impurities reagents, for which maximum impurities are listed. The Committee on Analytical Reagents of the ACS periodically publishes “Reagent Chemicals” listing specifications ( https://pubs.acs.org/isbn/9780841230460 ; accessed February 19, 2020). These reagent-grade chemicals are of very high purity and are recommended for quantitative or qualitative analyses.
Ultrapure reagents
Many analytical techniques require reagents whose purity exceeds the specifications of those described previously. Manufacturers offer selected chemicals that have been especially purified to meet specific requirements. There is no uniform designation for these chemicals and organic solvents. Terms such as spectrograde, nanograde, and HPLC pure have been used. Data of interest to the user (e.g., absorbance at a specific UV wavelength) are supplied with the reagent.
Other designations of chemical purity include chemically pure (CP), US Pharmacopeia (USP), and National Formulary (NF) grade (chemicals produced to meet specifications set down in the USP or the NF). Chemicals labeled purified, practical, technical, or commercial grade should not be used in clinical chemical analysis without previous purification.
Reference materials
A reference material is a material or substance with one or more physical or chemical properties that is sufficiently well established to be used for (1) calibrating instruments, (2) validating methods, (3) assigning values to materials, and (4) evaluating the comparability of results. Reference materials are of prime importance in establishing metrologic transferability ( http://www.bipm.org ; accessed February 19, 2020), , a term defined as “the property of a measurement result whereby the result can be related to a reference through a documented unbroken chain of calibrations, each contributing to the measurement uncertainty.”
For more detailed information on how reference materials are used in standardization and harmonization, see Chapter 7 .
Primary, secondary, standard, and certified are types of reference materials.
Primary reference materials
Primary reference materials are highly purified chemicals that are directly weighed or measured to produce a solution whose concentration is exactly known. The IUPAC has proposed a degree of 99.98% purity for primary reference materials. These highly purified chemicals may be weighed out directly for the preparation of solutions of selected concentration or for the calibration of solutions of unknown strength. They are supplied with a certificate of analysis for each lot.
Secondary reference materials
Secondary reference materials are solutions whose concentrations cannot be prepared by weighing the solute and dissolving a known amount into a volume of solution. The concentration of secondary reference materials is usually determined by analysis of an aliquot of the solution by an acceptable reference method, using a primary reference material to calibrate the method.
Standard reference materials
Standard reference materials (SRMs) for clinical and molecular laboratories are available from the National Institute of Standards and Technology (NIST; http://www.nist.gov ; accessed February 19, 2020). Cholesterol, the first SRM developed by the NIST, was issued in 1967. It should be noted that not all SRMs have the properties and the degree of purity specified for a primary standard, but each has been well characterized for certain chemical or physical properties and is issued with a certificate that gives results of the characterization. These may then be used to characterize other materials.
Examples of SRMs that are available from the NIST for use in clinical and molecular diagnostics laboratories include (1) pure crystalline standards ( Table 15.5 ), (2) human-based standards ( Table 15.6 ), (3) animal blood standards ( Table 15.7 ), (4) standards containing drugs of abuse in urine and human hair ( Table 15.8 ), and (5) SRMs used for DNA profiling/crime scene investigations ( Table 15.9 ).
TABLE 15.5
Standard Reference Materials: Pure Crystalline Standards
Available from the National Institute of Standards and Technology ( www.nist.gov ; accessed February 19, 2020).
| SRM Number | Analyte |
|---|---|
| 998 | Angiotensin I (human) |
| 916a | Bilirubin |
| 915b | Calcium carbonate |
| 911c | Cholesterol |
| 921 | Cortisol (hydroxycortisone) |
| 914a | Creatinine |
| 917c | D-Glucose (dextrose) |
| 920 | D-Mannitol |
| 937 | Iron metal (clinical) |
| 928 | Lead nitrate |
| 924a | Lithium carbonate |
| 929a | Magnesium gluconate dehydrate |
| 918b | Potassium chloride |
| 919b | Sodium chloride |
| 1595 | Tripalmitin |
| 912a | Urea |
| 913b | Uric acid |
| 925 | VMA (4-hydroxy-3-methoxy-DL-mandelic acid) |
| 8327 | Peptide reference materials (for molecular mass and purity measurements) |
SRM, Standard reference materials.
TABLE 15.6
Standard Reference Materials: Human Serum Based
Available from the National Institute of Standards and Technology ( www.nist.gov ; accessed February 19, 2020).
| SRM Number | Description |
|---|---|
| 909c | Human serum (contains 12 analytes) |
| 1951c | Lipids in frozen human serum |
| 956c | Electrolytes in frozen human serum |
| 965b | Glucose in frozen human serum |
| 967a | Creatinine in frozen human serum |
| 970 | Ascorbic acid in frozen human serum |
| 1952a | Cholesterol in human serum |
| 968e | Fat-soluble vitamins in human serum |
| 1589a | PCBs, pesticides, and dioxins/furans in human serum |
| 1599 | Anticonvulsant drug level assay (valproic acid and carbamazepine) |
| 900a | Antiepilepsy drug in frozen human serum |
| 1955 | Homocysteine and folate in human serum |
PCBs, Polychlorinated biphenyls; SRM, standard reference materials.
TABLE 15.7
Standard Reference Materials: Miscellaneous
Available from the National Institute of Standards and Technology ( www.nist.gov ; accessed February 19, 2020).
| SRM Number | Description |
|---|---|
| 955c | Toxic metals in caprine (goat) blood |
| 966 | Toxic metals in bovine blood |
| 1598a | Inorganic constituents in animal serum |
| 927e | Bovine serum albumin (7%, solution) |
| 2921 | Human cardiac troponin complex |
| 2389a | Amino acids in hydrochloride |
| 1400 | Bone ash |
| 1486 | Bone meal |
SRM, Standard reference materials.
TABLE 15.8
Standard Reference Materials for Drugs of Abuse in Urine and Human Hair
Available from the National Institute of Standards and Technology ( www.nist.gov ; accessed February 19, 2020).
| SRM | Description |
|---|---|
| 1508a | Cocaine metabolite in freeze-dried urine |
| 8444 | Cotinine in freeze-dried urine |
| 1507b | Marijuana metabolite in urine |
| 2381 | Morphine and codeine in urine |
| 2382 | Morphine glucuronide in urine |
| 1511 | Multiple drugs of abuse in urine |
| 2379 | Drugs of abuse in human hair I |
SRM, Standard reference materials.
TABLE 15.9
Standard Reference Materials for Use in DNA Profiling/Crime Scene Investigations
Available from the National Institute of Standards and Technology ( www.nist.gov accessed February 19, 2020).
| SRM | Description |
|---|---|
| 2372 | Human DNA quantitation standard |
| 2391c | PCR-based DNA profiling standard |
| 2392 | Human mitochondrial DNA sequencing (3 components) |
| 2392-1 | Human mitochondrial DNA sequencing (1 component) |
| 2394 | Heteroplasmic mitochondrial DNA mutation detection standard |
| 2396 | Oxidative DNA damage mass spectrometry standard |
PCR, Polymerase chain reaction; SRM, standard reference materials.
Certified reference materials
Certified reference materials (CRMs) are available for clinical and molecular laboratories from the Institute for Reference Materials and Measurements (IRMM) in Geel, Belgium ( https://ec.europa.eu/jrc/en/reference-materials ; accessed February 19, 2020). The IRMM is one of the seven institutes of the Joint Research Centre (JRC), a Directorate-General of the European Commission (EC). Other acronyms used to label IRMM reference materials include ERM (European Reference Materials), BCR (Community Bureau of Reference of the Commission of the European Communities), and the IFCC.
Examples of IRMM standards are listed in Tables 15.10 and 15.11 . Reference materials also are available from the World Health Organization (WHO; http://www.who.int/biologicals ; accessed February 19, 2020).
TABLE 15.10
Additional Certified Reference Materials
Available from the EU Science Hub ( https://ec.europa.eu/jrc/en/reference-materials ; accessed February 19, 2020).
| Number | Description |
|---|---|
| BCR-304 | Lyophilized human serum |
| BCR-573, 574, and 575 | Creatinine in human serum |
| IRMM-468 and 469 | Thyroxine (T 4 ) and triiodothyronine (T 3 ), two levels each |
| ERM-DA451/IFCC | Cortisol reference panel of fresh frozen human serum |
| ERM-DA192 and 193 | Cortisol in human serum |
| BCR-348R and BCR-DA347 | Progesterone in human serum |
| BCR-576, 577, and 578 | 17-β-estradiol in human serum |
| ERM-CE- 195 and 196 | Lead (Pb) and cadmium (Cd) in lyophilized bovine blood |
| BCR-634, 635, and 636 | Pb and Cd in lyophilized human blood |
| BCR-637, 638, and 639 | Aluminum, selenium, and zinc in human serum |
| BCR-393 | Human apolipoprotein A1 |
| BCR-457 | Human thyroglobin (Tg) |
| BCR-486 | Purified α-fetoprotein (AFP) |
| BCR-613 | Prostate specific antigen (PSA) lyophilized |
| BCR-405 | Glycated hemoglobin in human hemolysate |
| ERM-DA470k | Human serum proteins |
| ERM-DA474/IFCC | C-reactive protein (CRP) in frozen human serum |
| BCR-522 | Hemiglobincyanide (HCN) in bovine blood lysate |
| IRMM/IFCC-4467 | Hemoglobin isolated from whole blood |
| BCR-410 | Prostatic acid phosphatase from human prostate |
| BCR-647 | Human adenosine deaminase (ADA1) from human erythrocytes |
| BCR-693 | Human pancreatic lipase from pancreatic juice |
| BCR-694 | Human pancreatic lipase (recombinant) |
| ERM-AD452/IFCC | γ-Glutamyltransferase from pig kidney |
| ERM-AD454/IFCC | Alanine aminotransferase from pig heart |
| ERM-AD455/IFCC | Creatine kinase (CK-MB) from human heart |
| IRMM/IFCC-456 | Human pancreatic α-amylase |
| ERM-AD457/IFCC | Aspartate transaminase (AST) |
TABLE 15.11
Standards Certified for DNA Sequence a
Available from the EU Science Hub ( https://ec.europa.eu/jrc/en/reference-materials ; accessed February 19, 2020).
| Number | Plasmid DNA |
|---|---|
| IRMM/IFCC-490 | Sequence of 609-bp DNA fragment from human prothrombin gene (G20210 wild-type sequence) |
| IRMM/IFCC-491 | Sequence of 609-bp DNA fragment from human prothrombin gene (point mutation G20210A) |
| IRMM/IFCC-492 | Sequence of 609-bp DNA fragment from human prothrombin gene (G20210 wild-type and point mutation G20210A sequences) |
bp, Base pairs.
a Availability: each polypropylene vial contains approximately 1-ng plasmid DNA in a volume of 50 μL of a tris/ethylene diamine tetra-acetic acid solution.
Basic techniques and procedures
Basic practices used in clinical laboratories include (1) optical, (2) chromatographic, (3) electrochemical, (4) electrophoretic, (5) mass spectrometric, (6) enzymatic, and (7) immunoassay techniques. These techniques are discussed in detail in Chapter 16, Chapter 17, Chapter 18, Chapter 19, Chapter 20, Chapter 21, Chapter 22, Chapter 23, Chapter 24, Chapter 25, Chapter 26 . This chapter discusses the basic techniques of volumetric sampling and dispensing, centrifugation, measurement of radioactivity, gravimetry, thermometry, controlling hydrogen ion concentration, and processing solutions.
Volumetric sampling and dispensing
Clinical chemistry procedures require accurate volumetric measurements to ensure accurate results. For accurate work, only class A glassware should be used. Class A glassware is certified to conform to the specifications outlined in NIST circular C-602.
Pipettes
Pipettes are used for the transfer of a volume of liquid from one container to another. They are designed either (1) to contain a specific volume of liquid or (2) to deliver a specified volume. Pipettes used in clinical laboratories include (1) manual transfer and measuring pipettes, (2) micropipettes, and (3) electronic and mechanical pipetting devices. Developments in improved design of pipetting systems include robotic automation, the capability to provide electronic and personal computer control of pipetting devices, and careful attention to advanced ergonomic design features. Automatic photometric pipette calibration systems are also available that reduce the time needed to periodically check pipettes and potentially allow more efficient use of personnel.
Transfer and measuring pipettes.
A transfer pipette is designed to transfer a known volume of liquid. Measuring and serologic pipettes are scored in units such that any volume up to a maximum capacity is delivered.
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