(P) Lec 5: Analytical Techniques in CC (P2) Flashcards
1
Q
Refers to migration of charged particles in an electric field
A
Electrophoresis
2
Q
Separates proteins based on their electric charge densities
A
Electrophoresis
3
Q
Charge of proteins
A
Negative
4
Q
Buffer used in electrophoresis and its pH
A
Barbital/Veronal (pH = 8.6)
5
Q
Terms
Has a net charge of that can either be positive or negative depending on the pH condition
A
Amphoteric
6
Q
Terms
Movement of buffer ions and solvent relative to the fixed support (gel)
A
Electroendosmosis/Endosmosis
7
Q
Terms
Migration of small charged ions
A
Iontophoresis
8
Q
Terms
Migration of charged macromolecules
A
Zone Electrophoresis
9
Q
Familiarize the components of electrophoresis
A
- Electrical power
- Support medium
- Buffer
- Sample
- Detecting system
10
Q
What’s the detecting system of electrophoresis in MolBIo
A
UV light or radiophotography
11
Q
What are the factors affecting the rate of Migration?
A
Net electric charge of the molecule
Size and shape of the molecule
Electric field strength
Nature of the supporting medium
Temperature of the operation
12
Q
What are the supporting media in electrophoresis?
A
- Cellulose acetate
- Agarose gel
- Polyacrylamide gel
13
Q
Familiarize the stains for electrophoresis bands
A
Amido Black
Ponceau S
Oil Red O
Sudan Black
Fat Red 7B
Coomassie Blue
Gold/Silver Stain
14
Q
Visualization for separating lipoproteins with fats (lipids + proteins)
A
Oil Red O
Sudan Black
Fat Red 7B
15
Q
Staining protein found in CSF for staining during electrophoresis
A
Coomassie Blue
16
Q
Staining for smaller ions in electrophoresis
A
Gold/silver stain
17
Q
Measures electrophoretic bands
Measures the absorbance of the stain
Scans and quantitates electrophoretic pattern
A
Densitometry
18
Q
Types of electrophoresis
A
Isoelectric focusing & Capillary Electrophoresis
19
Q
Isoelectric electrophoresis separate by migration through what?
A
pH gradient
20
Q
Ideal for separating proteins of identical sizes but different net charges
Proteins move in electric field until they reach a pH equal to their isoelectric point
A
Isoelectric focusing
21
Q
Advantages of isoelectric electrophoresis
The ability to resolve mixture of _____ (identical in sizes)
Detects _______
identify genetic variants of ______
Detects CSF _______
A
protein
isoenzyme
genetic variants of proteins
oligoclonal banding
22
Q
In this type of electrophoresis, sample molecules are separated by Electro-Osmotic Flow
A
Capillary electrophoresis
23
Q
Capillary electrophoresis uses which unit to express quantity of specimen
A
Nanoliter
24
Q
Capillary Electro
Which charged ions in the specimen emerge early at the capillary outlet because of EOF and the ion movements are in the same directions?
A
Positively charged
25
Capillary Electro
Which charged ions in the specimen towards the capillary outlet but at a slower rate?
Negatively charged
26
Refers to separation of soluble components in a solution by specific differences in the physical and chemical characteristics of the different constituents
Chromatography
27
Familiarize basis of separation of chromatography
Rate of diffusion
Solubility of the solute
Nature of the solvent
Sample volatility/solubility
Distribution between 2 liquids
Molecular size
Hydrophobicity of molecule
Ionic attraction
Differential distribution between 2 immiscible liquids
Selective separation of substances
Differences in the absorption and desorption of solutes
28
Two forms of chromatography
Planar and column
29
Paper chromatography is used for the fractionation of which two components?
Sugar and amino acid
30
Refers to
Semi-quantitative drug screening test
Sample components are identified by comparison with standards on the same plate
Thin layer chromatography (TLC)
31
T or F: TLC is quantitative drug screening test
False (semi)
32
T or F: Extraction of drug in TLC is pH independent
False (pH dependent)
33
In TLC, this is defined as thin plastic plates impregnated with a layer of silica or alumina
Sorbent
34
Type of chromatography that uses a tube
Column
35
Used for the separation of steroids, barbiturates, blood, alcohol, and lipids
Gas chromatography
36
Useful for compounds that are naturally volatile or can be easily converted into a volatile form
Gas chromatography
37
Samples are introduced into the gas chromatography column using a _________________
hypodermic syringe or an automated sample
38
Elution order of volatiles in gas chromatography are based on what?
Boiling point
39
Mobile phase of samples in gas chromatography
Nitrogen
Helium
Hydrogen
Argon (Inert gasses)
40
In mass spectroscopy, the separation is based on what?
Fragmentation and ionization
41
MS can also detect _______ and determination of ______________
Structural information, molecular weight
42
T or F: Compound cannot be detected by MS if it isn't separated first by GC
True
43
Refers to double spectroscopy
Tandem Mass Spectroscopy
44
The tandem of GC and MS is called what?
Gas-Chromatography-Mass Spectroscopy
45
This can detect 20 inborn errors of metabolism from a single blood spot
Tandem Mass Spectroscopy
46
Gold standard for drug testing
GC-MS
47
Identify which is false about GC -MS
a.) Gold standard for drug testing
b.) Also used for xenobiotics, anabolic steroids, and pesticides
c.) Qualitative measurement of drug can be performed by selective ion monitoring
d.) Uses an electrode beam to split the drug emerging from the column into its component ions
C (guess why nalang)
48
GC-MS
The position of the parent-molecule ion and degradation products give rise to _____________ that which will provide the final identity of the drug of interest
Fingerprint patterns
49
Chromatography based on the distribution of solutes between a liquid phase and a stationary phase
Liquid chromatography
50
What is the fixed component and moving component in Liquid Chromatography?
Stationary phase: fixed component
Liquid phase: Moving component
51
This uses high pressure for fast separations, controlled temperature, in line detectors and gradient elution technique
High Performance Liquid Chromatography (HPLC)
52
Used in separation and quantification of various hemoglobin associated with specific diseases
HPLC
53
E2 eong c0nfiMatOrY miTod nG gc-ms, an0 i2????
koriqUe! liqUiD chRoma - Mas sPectRo
54
Also used in therapeutic drug monitoring, toxicology, and studies of drug metabolites
Requires interface methods to convert non-volatile to volatile compounds
LC-MS
55
What are the interface methods used in LC-MS?
Electrospray (ES)
Atmospheric Pressure Chemical Ionization (ARCI)
56
Familiar the separation mechanisms in Liquid Chroma
Gel/Gel Permeation/Gel Filtration/Size Exclusion/Molecular Sieve Chromatography
Ion exchange chromatography
Partition Chromatography
Affinity Chromatography
Adsorption Chromatography
57
determines the light emitted by a molecule after excitation by electromagnetic radiation
This is the principle of which mechanism/process?
FLUORIMETRY/MOLECULAR LUMINESCENCE
58
Light source of Fluorimetry/Molecular Luminescence
Mercury or Xenon Lamp
59
Light detector of Fluorimetry
Photomultiplier Tube or Phototube
60
This is used to measure porphyrins, magnesium, calcium, and catecholamines
Flourinetry
61
How many monochromators are used in fluorimetry?
Two
62
Assoc type, which is true about Fluorimetry
a. The incident light is prevented from striking the photodetector by the primary filter
b. The wavelength that is best absorbed by the solution to be measured is selected by the second filter
D (u rationalize it for more challenge, amen)
63
T or F: Fluorimetry is 100x more sensitive than spectrophotometer
False (1000x)
64
This is affected by quenching
Fluorimetry
65
Refers to the loss of fluorescence and radiation
Quenching
66
Familiarize the parameters affected by the quenching effect of fluorimetry
pH
Temperature changes
Chemical contaminants
UV light changes
67
Emission of light created by your sample after a chemical ог electrochemical reaction and not from absorption of electromagnetic energy
Chemiluminescence
68
The chemical reaction yields an electronically excited compound that emits light as it returns to its ground state or that it transfers its energy to another compound, which then produces emission
This is the principle of which mechanism/analytical technique?
Chemiluminescence
69
Use of chemiluminescence
Immunoassays (serology)
70
Photodetector of chemiluminescence
Photomultiplier tube (Luminometer)
71
Which is more sensitive between fluorescence and chemiluminescence?
Chemiluminescence
72
T or F:
In chemiluminescence, no excitation radiation is required and no monochromator are needed because the chemiluminescence arises from one species
True
73
Measurement of OSMOLALITY of an aqueous solution such as serum, plasma or urine
Osmometry
74
Based on measuring changes in the colligative properties of solutions that occur owing to variations in particle concentrations
This is the principle of which mechanism/analytical technique?
Osmometry
75
What are the four colligative properties involved in osmometry?
Osmotic pressure
Boiling point
Freezing point
Vapor pressure
76
These are the solutions measured in osmometry?
serum, plasma or urine
77
These are used on osmotic particles that when added, will increase the osmolality reading on the osmometer
Glucose
Urea
Sodium
78
Which of the colligative properties increases when osmolality is increased?
Osmotic pressure
Boiling point
79
Which of the colligative properties decreases when osmolality is increased?
Freezing point
Vapor pressure
80
Which is true about Freezing Point Depression Osmometry:
a.) Most commonly used method for measuring the changes in the colligative properties of a solution
b.) Based on the principle that addition of solute molecules lowers the temperature at which a solution freezes.
C
81
A 1.0 mOsm/kg solution has a freezing point depression at __________when compared with a pure solvent (usually water)
0.00186°C
82
Freezing point of abd osmolality of plasma?
Osmolality: 285 mOsm/kg
Freezing point: -53 ° C
83
What are measured in electrochemistry techniques?
Current and voltage
84
Electrochemistry
Measurement of electrical potential due to the activity of free ions
Follows the NERNST EQUATION
A change in the voltage indicates activity of each analyte
Potentiometry
85
Reference electrodes of potentiometry
Calomel and Silver chloride
86
Potentiometry uses which tests
pH and pCO2 tests
87
Electrochemical transducer capable of responding to one given ion
VERY SENSITIVE and SELECTIVE for the ion it measures.
Ion selective electrode
88
Ionic selectivity in Ion selective electrode depends on the ___________ used
membrane/barrier composition
89
T or F: Ion selective electron is selective, sensitive, and specific
False (not specific)
90
In Ion selective electrode, this measures the electrolytes dissolved in the fluid phase of the sample
ISE analyzers
91
ISE membrane
Glass membrane electrode measures what analyte?
Sodium
92
ISE membrane
Liquid membrane + valinomycin measures what analyte?
Potassium
93
ISE membrane
Silver membrane electrodes measures what analyte?
Chloride
94
T or F: Ion selective electrode does not discriminate between ions causing voltage differences between the measuring electrode and the standard electrode
Truew
95
Electrochemistry
Measurement of the amount of electricity in coulombs at a fixed potential
Electrochemical titration in which the titrant is electrochemically generated and the endpoint is detected at amperometry
Follows FARADAY's LAW
Coulometry
96
Test used in coulometry
Chloride test
97
Electrochemistry
Measurement of the current flow produced after by an oxidation reaction
Amperometry
98
Interferences in coulometry
Bromide
Cyanide`
Cysteine
99
Uses of amperometry
pO2
glucose
chloride
peroxidase determinations
100
Measurement of differences in current at a constant voltage
Follows the ILKOVIC EQUATION
Polarography
101
Electrochemistry
Measurement of current after which a potential is applied to an electrochemical cell
Allows the sample to be pre-concentrated, thus utilizing minimal analytes
Voltammetry
102
