Basic Principles and Practices of Clinical Chemistry Flashcards
What is a Primary Standard?
Highly purified sample with an exact known concentration
What is calibration?
Process that pairs an analytical signal (result from machine) with the concentration of an analyte
What is linearity studies?
The relationship between the concentration of standards and the instrument’s result
_______ establishes linearity
Calibration
What do you use to calibrate a machine?
Primary Standard
What type of water is standard in all labs?
Deionized Water
Define Solute
A substance dissolved in a liquid
Define Solvent
The liquid a substance is dissolved in
Define Solution
The combination of solute and solvent
Define Analyte
Substance of interest being measured
Define Percent Solution
Amount of solute per 100 total units of solution
Define Molarity
Number of moles per 1 L of solution
Define Molality
Amount of solute per 1 kg of solvent
Define Normality
The number of gram equivalent weights per 1 L of solution
What is the Henderson-Hasselbalch equation used for?
Is an equation that mathematically describes the dissociation characteristics of weak acids, bases, and its effect on pH
pH = pKa + log ([A-] / [HA])
Define Dilutions
The part(s) of concentrated material to the total final volume of a solution
Do you allow serum tubes to clot?
Yes
Define the light source, monochromator, Sample cell, and Photodetectors in Spectrophotometry
Light source provides light
Monochromator isolates wave lengths
Sample cell holds sample, maintains light path
Photodetector converts transmitted radiant energy into equivalent amount of electrical energy (absorbed vs transmitted)
What is Beer’s Law?
States that the concentration of a substance is directly proportional to the amount of light absorbed or inversely proportional to the logarithm of the transmitted light
Graphing Beer’s Law mathematically established the relationship between the analyte concentration and absorbance of light (linearity)
What are some components of Spectrophotometer Quality Assurance?
Wavelength Accuracy, Stray light, Linearity
What is transmittance?
The amount of light that passes through the sample and strikes the detector
What is absorbance?
Measure of light that is absorbed by sample
Define Atomic Absorption Spectrophotometry
What is special about Atomic Absorption Spectrophotometry?
Used to measure concentration by detecting the absorption of electromagnetic radiation by atoms rather than molecules
It is sensitive and precise. It it routinely used to measure the concentration of trace metals that are not easily excited
Define Fluorometry
What is its two advantages over conventional Spectrophotometry and one disadvantage?
Is part of Spectrophotometry (90 degree angle)
Measure the concentrations of solutions that contain fluorescing molecules.
Measurements are related to molar absorptivity of the compound
Measures UV light
Advantages: Specificity and Sensitivity
Disadvantage: very sensitive to environmental changes
Define Chemiluminescence
Is part of Spectrophotometry
Chemical energy generated produces excited intermediates that decay to a ground state with the emission of photons.
Used to measure enzymes
Define Turbidimetry
Is part of Spectrophotometry
Turbidimetry made with a spectrophotometer to determine the concentration of particulate matter in a sample.
Measures light blocked at 180 degree angle.
Define Nephelometry
Is part of Spectrophotometry
Nephelometry is similar to turbidometry, except that light scattered by the small particles is measured at an angle to the beam incident on the cuvette, instead of at 180°.
Define Spectrophotometry
Measures visible light
Measures light absorbance or transmittance from molecule(s)
Define Electrochemistry
The basis for many types of analyses used in the clinical laboratory, including potentiometry, amperometry, coulometry, and polarography
Molecules are made out of atoms…..we can use the atoms’ charges to measure the molecule
Define Ion Selective Electrodes (pH electrode)
Non-contamininating 2 electrodes into a salt bridge buffer
1 is a reference (or standard) electrode
1 is a indicator electrode
Measures liquid
“Gets dirty”
Unaffected by color or turbidity
Define Gas-Sensing Electrodes
- pCO2
- PO2
Measures gases
Enzyme Electrode, Colulometric Titration (sweat chloride), Anodic Stripping Voltammetry (lead testing)
Define Electrophoresis
What are the 5 components that it consists of?
Electrophoresis is the migration of charged solutes or particles in an electrical field.
Electrophoresis consists of five components: the driving force (electrical power), the support medium (gel), the buffer, the sample, and the detecting system.
Cellulose Acetate is routinely used for this method
(It is used for proteins)
Define Immunofixation
Is part of Electrophoresis
Used in the clinical laboratory to characterize monoclonal proteins in serum, urine, or cerebrospinal fluid (CSF)
Monoclonal proteins present will appear as a discrete band (with both a heavy and a light chain monospecific antiserum occurring at the same position).
Define Mass Spectrometry
Measures mass to charge ratio of molecular components. Definitive identification of samples eluting from GC or LC columns is possible when a mass spec-trophotometer (MS) is used as a detector.
Using mass to charge, to measure it
Charge = charge of ion
How can GC/MS, LC/MS, and Protemics of Mass Spectrometry be used in the Lab?
GC/MS systems are widely used for measuring drugs of abuse in urine toxicology confirmations.
LC/MS also has great potential for measuring low-level and mixed-polarity analytes such as vitamin D, testosterone, and immunosuppressant drugs due to its superior
Next generation of biomarkers for human diseases will be discovered using techniques found within the research fields of genomics and proteomics. (pathogen ID)
Define Osmometry
Osmometry is the principle of measuring the concentration of solute particles in a solution using one of the four colligative properties discussed below.
1. Osmotic pressure
2. Vapor pressure
3. Boiling point
4. Freezing point
What are some additional considerations for Automated Immunoassays (Immunoassay Basics)
An Antibody molecule recognizes and binds to an Antigen
.
Affinity refers to the strength of binding between a single binding site on the Antibody and its epitope.
Reagent Antibody production is achieved by polyclonal or monoclonal techniques.
What are some additional considerations for Automated Immunoassays (Turbidimetry and Nephelometry)
Turbidimetry (immunoturbidimetry) and nephelometry (immunonepholometry) are two related automated methods used to quantitate Ag–Ab complexes.
In general, turbidimetry is less sensitive than nephelometry.
What is a competitive immunoassay?
Labeled Antigen (Ag*) in the reagent competes with Antigen in the patient sample for a limited number of Antibody binding sites.
What is a noncompetitive immunoassay?
Also known as sandwich assays, use a labeled reagent Antibody to detect the Antigen. Excess labeled Antibody is required to ensure that the labeled Antibody reagent does not limit the reaction.
What are the steps in Automated Analysis?
- Specimen Preparation andIdentification
- Specimen Measurement andDelivery
- Open tube sampling (no cap)
- Closed tube sampling (cap piercer) - Reagent Systems and Delivery
- Chemical Reaction Phase
- Measurement Phase
- Signal Processing andDataHandling (manually electronic)
Pre-analytic Phase
Sample collection, identification, and processing
Preparation of the sample for analysis (centrifugation, dilutions, etc.), could use robotics.
Examination of sample integrity (good sample)
Analytic Phase
Dispensing of reagents and samples to testing site
Measuring of analytes of interest in a sample
Impacted by QC (established linearity) and calibration
Post-analytic Phase
Specimen storage
Data Management
- Reporting of results
What is Descriptive Statistics
Measures of Center, Spread, and Shape
Measures of Center
- Mean, Median, Mode
What is Standard Deviation?
Average distance from mean of data set to every value in data set (bell curve)
Measurement of Precision (Within-run vs Within-laboratory)
Within-run - repeatability (ex: repeat same result 30 times)
Within-lab, also referred to as across-run or between-run precision (ex: 4 machines have same result)
Define accuracy
How closely an instrument’s measured value of an analyte is to the true value of the analyte
(within range or not)
Analytic Sensitivity
The ability of a method to detect small quantities or changes in concentration of an analyte
Analytic Specificity (Cross-Reactivity)
Ability of a method to detect only the analyte of interest
Diagnostic Sensitivity
The ability of a test to detect a given disease or condition
Diagnostic Specificity
The ability of a test to correctly identify the absence of a given disease or condition
Random error
Something you/environment does. Can be positive or negative
Systemic error
Error of system like systems or reagents (system not being recalibrated….over time, error increases)
What is the operation of a QC system?
- Establishing or verifying allowable statistical limits of variation for each analytic method
- Using these limits as criteria for evaluating the QC data generated for each test
- Taking action to remedy errors real time when indicated
- Finding the cause(s) of error
- Taking corrective action
- Reanalyzing control and patient data
What is proficiency testing?
The laboratory must incorporate proficiency testing into its routine workflow as much as possible.
Proficiency samples are tested by bench technical staff who normally conduct patient testing
Testing should be completed within the usual time it would take for routine patient testing.
Proficiency samples are to be performed and submitted on the primary analyzer
At least twice per year, other analyzers within the laboratory are to be compared to the primary analyzer.
All proficiency failures and significant shifts and trends must be reviewed, investigated, and resolved within 30 days of final receipt of proficiency results.
What reagent/chemical is used for lab use?
Reagent grade or analytic reagent (AR)
