Analytical Methods Flashcards
principle: measurement of light transmitted by solution to determine concentration of light-absorbing substances in solution
identify the analytical method
spectrophotometry
describe double beam spectrophotometer. how does it work?
monochromatic light split into two components. absorbance of sample can be recorded directly
State Beer’s Law
concentration of a substance is directly proportional to the light absorbed, or inversely proportional to the logarithm of transmitted light
purpose of collimator @ spectrophotometer
minimizes unwanted/scattered light
purpose of monochromator @ spectrophotometer
wavelength selector
T/F
Glass cuvette is used for both visible and UV light
F
quartz: UV radiation
identify:
convert transmitted radiant energy into equiv amt of electrical energy
photodetector
Amount of light absorbed at wavelength depends on molecular and ion types. Which 3 factors may affect?
concentration
pH
temperature
Differentiate reagent blank fr sample blank
reagent blank: corrects absorbance caused by color of reagent
sample blank: corrects absorbance caused by optical interference
identify instrument: measures absorption of light of unique wavelength by atoms @ ground state
atomic absorption spectrophotometer
Light source for Atomic absorption spectrophotometer
Hollow cathode lamp
Which modulates light source @ atomic absorption spectrophotometer? A. Chopper B. Nebulizer C. Both A and B D. NOTA
A
Which converts ions @ atomic absorption spectrophotometer? A. Chopper B. Nebulizer C. Both A and B D. NOTA
B
atomic absorption spectrophotometer applied at the ff except- A. Electrolytes B. Dissolved gases C. Carboxylic Acids D. NOTA
C
Before mass spectrometry, the compound must be isolated via gas chrom/HPLC.
True or false?
True
Mass spectrometry is used in: A. Drug testing B. Proteomics C. Both A and B D.NOTA
C
Identify analytical method
Measures amt of light emitted by a molecule after excitation by EM radiation
Fluorometry
[Fluorometry]
Purpose of primary filter @ monochromator
Allow passage of light of the proper wavelength for absorption of molecule
Factors that affect fluorescence include all except A. pH changes B. Temperature C. Exposure duration D. Hue of substance
D
Concentration dapat
False about chemiluminescence
A. Frm species
B. Emitted radiation is related to analyte concentration
C. Chemical rxn: electronically excited cpd that emits light as it returns to ground state
D. Signal is taken as derivative of entire peak
D
integral dapat
Main disadvantage of chemiluminescence
Impurities can cause background signal that degrades sensitivity and specificity
Identify analytical method.
Measurement of light scattered by particulate (turbid solution). Light has signal that must be amplified
Nephelometry
[Nephelometry] Light scattering dep on all except A.wavelength of incident light B.particle size C.amount of sample
C
[Nephelometry]
wavelength of incident light __ particle diam, para at least forward
less than or equal to
[Nephelometry]
Main application
measurement of Ag-Ab complex
[Nephelometry]
Identify type: wavelengths use are 320-650nm. Light scattered forward
Rayleigh-Debye type
[Nephelometry]
T/F
Detector output is inversely prop to concentration
F
Direct prop dapat
Measures reduction in light transmission due to particle formation
Identify analytical method
[Turbidimetry]
[Turbidimetry]
Amount of light absorbed by a suspension of particles depends on two factors: __ & __
Specimen conc
Particle size
Turbidimetry may be applied in all except A. Glucose measurement of CSF B. Detect bacterial growth in broth C. Detect clot formation D. Protein measurement in urine
A
[Electrophoresis]
The ff affect rate of migration except A. Shape of molecule B. Electric field strength C. Nature of supporting medium D. Pressure applied to machine
D
Temp dapat
[Electrophoresis]
The ff separate by molecular size except
A.Starch gel
B.Agarose gel
C.Polyacrylamide gel
B
[Electrophoresis]
Which supp medium does not bind protein?
A.Starch gel
B.Agarose gel
C.Polyacrylamide gel
B
Identify the analytical method:
charged proteins migrate through support medium that has continuous pH gradient
isoelectric focusing
isoelectric focusing: applied in two ways. enumerate
- detect oligoclonal immunoglobulin bands in CSF
2. detect enzymes like ALP in serum
identify instrument:
measures absorbance of the stain on a support medium
densitometer
[Densitometry]
Signals detected by photodetector: related to absorbance of sample stain on the support. This is proportional to ___
specimen concentration
identify analytical method:
measurement of electrical potential due to activity of free ions, or differences in voltage at constant currrent
potentiometry
[potentiometry]
activity of analyte indicated in __
change in voltage
[potentiometry]
applied in: __ and __
electrolytes
Dissolved gases
identify analytical method:
measurement of the current flow produced by redox reaction
amperometry
identify analytical method:
method in which potential is applied to electrochemical cell, and resulting current is measured
voltametry
identify analytical method:
measurement of quantity of electricity needed to convert analyte to diff oxid state.
coulometry
coulometry: direct application?
used to measure chloride ion in serum, plasma, CSF & sweat samples
identify analytical method:
separation method based on diff interactions of specimen compounds with mobile phase and stationary phase, as compounds travel thru support medium
chromatography
chromatography methods useful for compounds that are naturally inclined to be volatile; used in separation of blood, alcohol, lipids
gas chromatography
Gel chromatography separates molecules based on size and shape. It may use hydrophilic gel or hydrophobic gel. Uses of the former?
hydrophilic gel: sep antibodies, enzymes, proteins
Gel chromatography separates molecules based on size and shape. It may use hydrophilic gel or hydrophobic gel. Uses of the latter?
hydrophobic gel: sep triglycerides, fatty acids
identify chrom method
exchange of sample ions and mobile-phase ions with charged grp of stationary phase
ion exchange chromatography
identify chrom method
separation of compounds is based on partition bet liquid mobile phase and liq stationary phase, coated on solid support
partition chromatography
identify chrom method
uses immobilized biochem ligands as stationary phase to separate a few solutes fr other unretained solutes
affinity chromatography
identify chrom method
separation based on absorption/desorption of solutes @ surface of solid particle
adsorption chromatography
affinity chromatography may separate all except A. lipoproteins B. carbohydrates C. glycated hemoglobins D. vitamins
D
Enumerate 4 types of analytic errors
random
systematic
total
idiosyncratic
Enumerate 4 advantages of automating lab testing
- eliminate repetitive stress injury
- less error rates
- reduce operator occup hazard
- increase quality of pre-analytic steps
Enumerate 3 sources of problems, in sample introduction @ analytic stage
a. formation of clot attached to probe
b. inadequate or short sample
c. carry over
