clin chem exam 1 (unit 1)

therapeutic range

ref range used for drugs levels

reference range

“normal”, expected values for healthy individual, used to diagnose or monitor conditions, based on each labs population (age, sex, excludes pregnant ill tobacco users from pop), middle 95% reference sample in graph (right outside range might not indicate issue)

analytic sensitivity

probability of detecting all patients with the target disease, 100% = NO FALSE NEGATIVE, Smoke detector easily triggered by candle, if highly sensitive negative result rules out target disease SNOUT

diagnostic sensititvity

Ability of a test to detect a given disease or condition, positive means present

analytic specificity

probability of detecting only target disease, 100% = NO FALSE POSITIVE, higher this lowers sesitivity, smoke detector triggered by candle is not specific for house fire, if highly specific positive result rules in target disease SPIN

diagnostic spec

Ability of a test to correctly identify the absence of a given disease or condition, negative rules out disease

negative prediction value

dependant on population, probability that negative result = patient does not have the disease, 100% = NO FALSE NEGATIVE, Neg flu in summer is more believable (higher NPV) than a neg in winter (lower NPV), higher for rare condition, lower for common diseases

positive prediction value

dependent on population, probability that positive result = patient has the disease, 100% = NO FALSE POSITIVE, lower for a rare condition, higher for common disease

QA programs

preventative, assess the effectiveness of the lab's policies and procedures, identify and connect problems, assure the accuracy of reporting of results, assure competency of staff

calibration

Calibrator (standard)- most accurate material with known value to set analyzers to detect value and range, establishes AMR, done after instrument installation, using a new lot of reagent, to set regular intervals determined by test system/reagent manufacturer, to recalibrate following QC shift or trend

linear range = analytical measurement range (AMR)

the range determined by manufacturers and confirmed by labs 2x a year of results, instrument response and specimen value is linear, limitation of analyzer / reagent, can’t report results outside linear range

clinically reportable range (CRR)

also called maximum dilution or extended AMR, extended with a dilution of the sample, Kidney disease causes protein in the urine so doctors want to know extended dilutions of this

QC

assess quality of analysis methods (analyzers, employees, specimen conditions, etc…), measures accuracy and precision, check for random and systematic error

accuracy vs precision

a- close to correct, p- repeatedly getting same result

QC material/ matrix

similar to actual specimen, human or animal (biohaz), ideally stable forever (some can only be thawed so many times so aliquot and thaw as needed), normal and abnormal ranges (2-3 levels), lyophilized (freeze dried, reconstitution cause error, requires accurate pipetting), liquid pooled frozen pooled

assayed vs unassayed

a- tested many times before sold, gives specific target and range, expensive

u- lab must test to find target and range before using, 30x over 30 days by diff techs, broader less useful range

target value or mean

SD (spread of values around the mean, the higher the wider or further from 0 the variation and less precise), CV coefficient of variation (SD/mean x 100, used to compare different test’s SDs)

causes of trends and shifts

t- reagent deterioration and the light source deterioration

s- change in calibration reagent lot, improper reconstitutions, analyzer issues

random error issues

technique, use of non reagent grade water, incorrect reconstitution of control material, power supply, pipetting mistakes, analyzer pipette problem, air bubbles in sample

troubleshooting random error

re-assay fresh aliquot of same control material, open or reconstitute new vial of control material and re-assay

systemic error

improper alignment of sample or reagent pipettes, unstable incubator chamber, change of reagent lot or calibrator lot, deterioration of reagents or control material in use or light source, evaporation of sample during analysis

troubleshooting systemic error

check if reagent level is low and replace, resolve alarms or call tech support, if error on 2 diff analyzers indicates problem with QC material or something

last resort troubleshooting QC

recalibrate, consult supervisor or tech support, replace reagent pack

Given the following Level I control results for glucose, what remedial action should be taken based on the Day 10 result? day 1-9 ranged from high 95-107 by day 10 dropped to 89

Rerun the same Level I control.

westgard multirules

1 2S- in control, 1 point outside 2SD, warning, random error

1 3S- 1 point outside 3SD, random

R 4S- 1 point outside of 2 SD and another -2SD within same run, (2+2=4), random error

2 2S- 2 points outside of 2SS, systematic error

4 1S- 4 points outside 1SD, systemic

10x- 10 points on one side of mean/target, systemic

delta checks

compares patients current result with most recent previous result, flags changes greater than threshold, could indicate IV contamination, dilution/concentrat, treatment, wrong patient, instrument errpr

waved test

can be preformed without training or professional

proficiency testing (surveys)

external QC, CLIA requires for moderate-high complex testing, labs report back to CAP and API results from controls they send, should match other labs results

spectrophotometry and turbidimetry measures

light absorbed by analyte by measuring what is transmitted (light out / light in or transmitted / incident light)

fluorometry and chemiluminescence measure

light emitted by analyte due to photon energy

basic fluorometry

sample absorbs excited light, electron jumps to excited state, elections returns to ground emitting light proportional to analyte concentration, obsolete (sensitive to envir changes), 90 degree angle to avoid measuring the excitation light

fluorometry process

light source (LED or laser)> filters before and after sample> cuvette> photomultiplier tube

obsolete, therpeutic drugs

chemiluminescence

chemical reaction causing emission of light (luminescence), involves the oxidation of a luminecent substrate (luminal, acridinium esters, dioxetanes)

chemiluminescence process

light source (none)> filters> reactive cuvette (magnetic bead)> photomultiplier tube

roche cobas immunoassay, troponin, TSH, hCG

nephelometry measures

light scattered by analyte due to particles suspended in solution, 90 degree angle with light, immune complexes increase scattered light, increase in scatt is proportional to analyte concentration

nephelometry process

light source (laser)> filters> cuvette> photodiode or photomultiplier tube

siemens BN II system, IgG, IgA, C3, C4

light source

provides light of appropriate wavelength for the target analyte, tungsten, deuterium, halogen, LED

monochromator

selects a narrow wavelength bandpass to improve specificity for target analyte, measures single wave length to give signal-noise ration, simple colored glass filter, prism, diffraction grating (reflective surface with fine grooves separating light)

sample holder

flow cell, cuvette

photodetector

converts light into electrical signal, largely obsolete (photocell, phototube), photomultiplier tube PMT (amplifies small signal with dynodes), photodiode array PDA (measures multiple wavelength at once)

signal processing

converts signal to result

photometry

measures reflection of light intensity, beckman coulter iris, urine gllucose, protein, leukocytes

light source (LED)> filters> test strip> photodiode> result

simple reflection- visible light that isn’t absorbed is reflected

complex spectrophotometry- tells how much an analyte absorbs light

Beers Law

absorbance A is directly proportional to concentration c and path length (b, length of light path through sample cuvette, constant), a= molar absorptivity for each analyte, constant

A=abc

C/A = unknown C/A

transmittance is inversely proportionate to concentratuin

spectrophotometry process

light source (tung-hal VIS or deu UV)> diffraction grating> cuvette> photodiode or PDA

roche cobas chem, glucose, AST, creatinine

turbidometry

spectrophotometry measuring decrease in transmitted light, not translucent, immune complexes form and increase sample turb, turb increase with trans decrease is proportional to analyte concentration, detector is in line with light

turbidometry process

light source (tung-hal VIS or deu UV or xenon UV visable)> diffraction grating> cuvette> photodiode

roche cobas chem, CRP, HbA1c

Ion-selective Electrode (ISE)

measure the potential (voltage) between two electrodes in a solution, changes due to analyte concentartaion, used for electrolytes (sodium, potassium, chloride)

visible light

400-700

concentration =

peak height/ area

electrophoresis

use of an electric field to separate charged molecules based on charge, size, and shape, anode + cathode - (PANIC), electric fields, support medium, buffer, sample, detecting system

immunofixation

electrophoresis that uses ab to identify protein present in bands

capillary

electrophoresis that

isoelectric focusing

electrophoresis with pH gradient to separate proteins by their isoelectric point

2D

electrophoresis that

chromatography

mobile phase + stationary phase, compounds that react more strongly stauinary are retained longer, like dissolves like, retention time (how long each analyte take to elute off), resolution (how well the system separates compounds in sample)

absorption

chromatography separates by binding strength to solid surface, liquid mobile phase + solid surface staionary phase, analytes more faster in the mobile phase

partition

chromatography separates by mixing polar (aqueous) liquid phase with nonpolar (organic) liquid phase

ion exchange

chromatography separates by mobile phase similarly charged particles flowing through and opposite charge particles stick to the surface of stationary phase, bind and elute, flow-through

thin-layer chromatography TLC

mostly obsolete, retention factor Rf= distance of comp / total distance of solvent

high performance liquid chromatography HPLC

therapeutic drug monitoring, toxicology

gas chromatograohy GC

therapeutic drug monitoring and toxicology

mass spectrometry

identify and quantify molecules in a sample based on mass-to-charge ratio, sample is vaporized and ionized (electrospray or matrix- assisted laser desorption ionization MALDI), and passes through strong magnetic field to separated charged particles allowing ions to reach detector at different rates