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Analytical Methods Flashcards

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1
Q

Energy are electromagnetic waves that are characterized by ___________ and ___________.

A

Frequency and Wavelength

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2
Q

Wavelength

A

HORIZONTAL (width) distance from each wave

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3
Q

Amplitude

A

VERTICAL (height) distance from each wave

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4
Q

Frequency

A

Number of waves/vibrations

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5
Q

Wavelength is ___________ proportional to Frequency and Energy

A

Inversely proportional

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6
Q

Frequency is _________ proportional to Energy

A

Directly Proportional

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7
Q

Wavelengths are expressed in _________.

A

Nanometer

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8
Q

Increased Color Reaction: ________________

A

Increased analyte concentration

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9
Q

Colorimetric Methods

A

Spectrophotometry
Flame Emission Photometry
Atomic absorption Spectrophotometry

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10
Q

Principle in Spectrophotometry

A

Measurement of the LIGHT TRANSMITTED in the solution to determine the concentration of the LIGHT ABSORBED in the solution

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11
Q

______________ is used to verify WAVELENGTH ACCURACY and for instruments with narrower bandpass.

A

Mercury Vapor Lamp

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12
Q

Light Source

A

a.k.a Radiant energy source/Lamps
Provide limited radiation and polychromatic light

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13
Q

Most common light source for VISIBLE-INFRARED light

A

Tungesten

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14
Q

Routinely used for UV light

A

Deuterium

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15
Q

Enumerate: Light Source (UV Light)

Mnemonics: MXHD

A

Mercury Arc
Xenon
Hydrogen Lamp
Deuterium

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16
Q

Enumerate: Light Source (Infrared Light)

Mnemonics: MG

A

Merst Glower
Globar

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17
Q

Number of Photodetectors in Double Beam in SPACE

A

2 photodetectors

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18
Q

Number of Photodetectors in Double Beam in TIME

A

1 photodetector

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19
Q

SIMPLEST type of absorption spectrophotometry

A

Single Beam spectrophotometry

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20
Q

Single Beam Spectrophotometry

A

One measurement at a time

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21
Q

Double Beam Spectrophotometry

A

Splits monochromator light into 2 components

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22
Q

Emits radiation that CHANGES INTENSITY

A

Continuum Source

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23
Q

Emits limited radiation in discrete lines

A

Line Source

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24
Q

Monochromator

A

“Wavelength Selector” - Isolate specific or individual light

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25
Q

Most common and Best utilized monochromator

A

Diffraction Gratings

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26
Q

Synonyms of “Cuvette”

A

Sample Holder
Absorption Cell
Analytical Cell
Sample Cell

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27
Q

Cuvette

A

Holds the solution which concentration is to be measured

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28
Q

Most Common sample holder

A

Aluminum Silica Glass

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29
Q

Sample holder BEST utilized for UV

A

Quartz

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30
Q

Photodetector

A

Detect and Convert transmitted ight into PHOTOELECTRIC ENERGY

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31
Q

Most common Photodetector

A

Photomultiplier Tube (PMT)

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32
Q

Excellent in LINEARITY

A

Photodiode

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33
Q

Routine path length of of Cuvettes

A

1 cm

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34
Q

Range of Aluminum Silica Glass cuvettes to transmit light effectively

A

> 220 nm

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35
Q

What to do when results are NON-LINEAR?

A
  1. Blank Technique
  2. Reagent Blank
  3. Sample Blank
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36
Q

Most common cause of loss of Linearity

A

Stray Lights

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37
Q

Principle in Flame Emission Spectrophotometry (FEP)

A

Light Emitted from EXCITATION of electrons from lower to higher energy source.

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38
Q

Flame Emission Photometry measures light emitted by a ___________ burned in a ________.

A

Single atom
Flame

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39
Q

FEP.

Light Source: ___________
Photodetector: _______

A

Light Source: Flame
Photodetector: Photocell

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40
Q

Indirect Internal standard Method (Recalls: March, 2019)

Sodium: ________
Potassium: _________
Calcium: __________
Lithium: __________

A

Sodium: Yellow
Potassium: Purple
Calcium: Orange
Lithium: Red

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41
Q

Quality Assurance in FEP. Flickering Lights indicates changes in the ____________ of the instrument.

A

Fuel reading

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42
Q

Preferred Internal Standard Method (FEP)

A

Lithium
Cesium

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43
Q

Principle: Atomic Absorption Spectrophotometry

A

Elements are not excited by merely dissociating from its chemical bond and are placed in an unexcited, unionized, ground state.

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44
Q

Atomic Absorption Spectrophotometry (AAS)

Light Source: ________
Photodetector: _________

A

Light Source: Hallow Cathode Lamp
Photodetector: PMT

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45
Q

Reference method for CALCIUM, MAGNESIUM, TRACE METALS.

A

Atomic Absorption Spectrophotometry

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46
Q

Principle: Volumetry/Titrimetry

A

Unknown sample is made to react with Known sample in the presence of INDICATOR

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47
Q

Methods using Volumetry

A

Schales and Schales: Chloride
EDTA Titration Method: Calcium

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48
Q

Principle: Turbidimetry

A

Measures the amount of light BLOCKED in a turbid solution.

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49
Q

Turbidimetry measures abundant _________ like proteins and bacteria.

A

Large Particles

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50
Q

Principle: Nephelometry

A

Measures the amount of Light SCATTERED in a turbid solution.

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51
Q

Analytical method used to measure antigen-antibody complex

A

Nephelometry

52
Q

Measure angle from scattered light in cuvet

A

15 - 90 degrees

53
Q

Principle: Fluorometry

A

Measures the amount of light emitted by a MOLECULE after EXCITATION through ELECTROMAGNETIC RADIATION.

54
Q

Only method that uses 2 Monochromators

A

Fluorometry

55
Q

Fluorometry measures light intensity in a _______ background.

A

Zero background

56
Q

Purpose of 2 Monochromators in Fluorometry

A

1st - Primary/Excitation Filter: selects WL that is best absorbed
2nd - Secondary/Emission Filter: prevent incident light

57
Q

Fluorometry

Light Source: ________
Photodetector: ________

A

Light Source: Mercury, Xenon, UV light
Photodetector: PMT/Phototube

58
Q

What is Quenching?

A

Reduction or limitation of a particle in an excited state through;

pH changes
Temperatures
UV
Chemical changes

59
Q

Recalls: September 2019

Increase Light: ______
Increase Temperature: _______
Increase QUENCHING: ________
Increased Absorbing Analyte: _______

A

Increase Light: Increase Fluorescence
Increase Temperature: Decrease Fluorescence
Increase QUENCHING: Decrease Fluorescence
Increased Absorbing Analyte: Decrease Fluorescence

60
Q

Scoring of FITC: Apple green Fluorescence

A

2+

61
Q

Difference of Maximum WL of excitation and Emitted Fluorescence or Max. WL of excitation - Emitted Fluorescence

A

Stokes Shift

62
Q

Auramine Rhodamine: _________
Calcofluor White: ________
Acridine Orange: __________

A

Auramine Rhodamine: Mycobacterium
Calcofluor White: Fungi and Yeasts
Acridine Orange: DNA - nucleus, nucleoid, plasmids.

63
Q

Principle: Chemiluminescence

A

Chemical/Electrochemical Reaction that yields an electronically EXCITED COMPOUND that emits light as it returns to its ground state.

64
Q

Chemiluminescence ________ use Light source and Monochromator

A

DO NOT use light Source and Monochromator

65
Q

Which is not part of Chemiluminescence?

  1. Light Source
  2. Photodetector
  3. Monochromator
  4. Cuvette
A

Light Source and Monochromator

Note: If there is BOTH Light Source and Monochromator are present in the choices, then Both is the answer. If BOTH is none from the choices, answer Monochromator only.

66
Q

Main Components of Luminometer

A

Cuvette
Photodetector
Read-out device.

67
Q

Principle: Osmometry

A

Measure of changes in Colligative Properties due to variation in solute/particle concentration.

68
Q

Colligative Properties

A

Osmotic Pressure
Boiling Point
Freezing Point
Vapor Pressure

69
Q

Increased Osmolarity (Recalls. September 2019, January 2021)

Increased - ________, _________
Decreased - __________, ________

A

Increased: Osmotic Pressure, Boiling Point

Decreased: Freezing Point, Vapor Pressure

70
Q

Principle: Denisitometry

A

Measures the concentration of ABSORBED stain from PROTEIN FRACTIONS

71
Q

Unit of Densitometry

A

Optical Density (Absorption)

72
Q

OD 450

A

Cut-off detection limit - BILIRUBIN

73
Q

OD 650

A

Cut-off detection limit - LAMELLAR BODIES

74
Q

Principle: Chromatograhy

A

SEPARATION of soluble materials in a solution by their CHEMICAL and PHYSICAL differences.

75
Q

Factors affecting Chromatraphy

A

Rate of Diffusion
Solubility
Size
Ionic Charges

76
Q

Planar Form Chromatography: __________
Column Form Chromatography: __________

A

Planar Form Chromatography: diffusion of molecules in 2D PLANE SYSTEM
Column Form Chromatography: diffusion of molecules in 3D MULTIDIRECTIONAL SYSTEM

77
Q

Paper Chromatography

Fractionation of _________ and _________.
Sorbent: ____________

A

Fractions: Sugar and Amino Acid

Sorbent: Whattman paper

78
Q

Thin Layer Chromatography

Use: _____________
Sorbent: _____________

A

Use: TDM and SEMI-QUANTITATIVE DRUG SCREENING TEST
Sorbent: Thin Plastic plate (Impregnated with a layer of Silica gel/Alumina)

79
Q

Used for VOLATILE compounds

A

Gas Chromatography

80
Q

Gas Chromatography separates ____________, __________, ___________, __________.

Mnemonics: LABS

A

Lipids
Alcohol
Barbiturates
Steroids

81
Q

Absorption of gases at SOLID phase surfaces

A

Gas Solid Chromatography

82
Q

Gas liquid chromatography is the partition between gaseous _________ phase, and liquid ________ phase.

A

Gaseous Mobile Phase and Liquid Stationary Phase

83
Q

Liquid Chromatography

A

Distribution of solute between liquid mobile phase and liquid stationary phase.

84
Q

Most widely used Liquid Chromatography

A

High Performance Liquid Chromatography (HPLC)

85
Q

Separate non-volatile substances

A

Liquid Chromatography - Mass Spectroscopy (LC-MS)

86
Q

Principle of Mass Spectrophotometry

A

Fragmentation and Ionization of molecules

87
Q

Gold Standard for Drug Testing

A

GC-MS

88
Q

Gold Standard for New Born Screening

A

MS-MS

89
Q

Migration based on ELECTRICAL CHARGE

A

Electrophoresis

90
Q

Migration based on PHYSICAL and CHEMICAL properties

A

Chromatography

91
Q

Migration based on pH GRADIENT

A

Isoelectric focusing

92
Q

Migration based on ELECTRO-OSMOTIC FLOW (EOF)

A

Capillary Electrophoresis

93
Q

Isoelectric Point

A

pH at which substances have a ZERO NET CHARGE (positively charged groups are equal to negatively charged groups)

94
Q

Electro-osmotic flow (EOF)

A

A bulk flow resulting from excess positive ions at the inner capillary surface moving towards the cathode

95
Q

Principle: Potentiometry

A

Measures electrical potential due to its activity of free ions

96
Q

Change in Voltage: _______________

A

Change in Voltage: Analyte Activity

97
Q

Potentiometry follows __________ equation

A

NERNST Equation

98
Q

Potentiometry measures _______ and _______.

A

pH and pCO2

99
Q

Two Types of ISE

A

Direct ISE - Undiluted Sample
Indirect ISE - Diluted Sample

100
Q

ISE Membranes

Glass Aluminum Silicate: _________
Valinomycin: ____________
Organic Liquid: __________
Glass Membrane: _________

A

Glass Aluminum Silicate: Sodium
Valinomycin: Potassium
Organic Liquid: Calcium, Lithium
Glass Membrane: pH

101
Q

ISE Electrodes

Clarke:
Severinghaus:
Glass Aluminum Silicate:
Valinomycin Gel:
Glass Membrane:

A

Clarke: pO2
Severinghaus: pCO2
Glass Aluminum Silicate: Na
Valinomycin Gel: K
Glass Membrane: pH

102
Q

Principle: Coulometry

A

Measures electricity at FIXED POTENTIAL

103
Q

The endpoint in Coulometry is detected by __________.

A

Amperometry

104
Q

Measured by Coulometry

A

Chloride Tests - CSF, Sweat, Serum

105
Q

Coulometry follows _________ Law

A

Faraday’s Law

106
Q

Measured by Amperometry

Mnemonics: PoGI C Papi

A

pO2
Glucose
Chloride
Peroxidase

107
Q

Measurement of differences in current at a constant voltage

A

Polarography

108
Q

Polarography uses __________ equation

A

Ilkovic Equation

109
Q

Nernst equation: ___________
Faraday’s Law: ___________
Ilkovic Equation: __________

A

Nernst equation: Potentiometry
Faraday’s Law: Coulometry
Ilkovic Equation: Polarography`

110
Q

Principle: Voltametry

A

Measurement of CURRENT AFTER A POTENTIAL IS APPLIED to an ELECTROCHEMICAL CELL.

111
Q

Anodic Striping Voltametry is utilized for _______, and ________ studies.

A

Lead and Iron

112
Q

Gene Mutation in Cystic Fibrosis

A

Chromosome 7

113
Q

Cystic Fibrosis (MUCOVISCIDOSIS)

Defect:
Sweat Inducer:
Diagnostic Test:
Reference Method:

A

Defect: Trans-epithelial Chloride Transport
Sweat Inducer: Pilocarpine
Diagnostic Test: Sweat Test Coulometry
Reference Method: Gibson and Cooke Pilocarpine Iontophoresisi

114
Q

Most Common

UV Lamp:
Visible - Infrared Lamp:
Cuvet:
Monochromator:
Photodetector:

A

Most Common

UV Lamp: Deuterium
Visible - Infrared Lamp: Tungsten
Cuvette: Aluminum Silica Glass
Monochromator: Diffraction Gratings
Photodetector: PMT

115
Q

Most sensitive and specific method in Colorimetry

A

AAS

116
Q

Quality assurance of Spectrophotometry

A

Wavelength Accuracy
Stray Light
Linearity

117
Q

PMT is _____ times more sensitive than phototube

A

200x

118
Q

Photocell vs. Phototube

A

Photocell - no external voltage
Phototube - require external voltage

119
Q

Stray Light

Causes:
Major Effect:
Detected:

A

: light outside the band by the monochromator

Causes: Scratches, Dust
Major Effect: absorbance error
Detected: Cutoff Filters

120
Q

Reference Method for Ca and Mg

A

AAS

121
Q

Modulate the light source in AAS

A

Mechanical Rotating Chopper

122
Q

Degree of Sensitivity of Analytical Method

A

Chemiluminescence > Fluorometry > Spectrophotometry

123
Q

Advantages of Chemiluminescence

A

Subpicomolar detection limits
Speed (Flash type rxns. & Light measured within 10 secs.)
Ease of use
Simple Instrumentation

124
Q

This technique contains the serum but without the reagent

A

Blanking Technique

125
Q

Interferes mainly in blanking technique, thus increasing light scatter (turbidity)

A

Lipids

126
Q

Verify Absorbance accuracy on Linearity

A

Neutral Density
Dichromate Soln

Clinical Chemistry (20 decks)

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