Electrophoresis Flashcards
1
Q
It helps establish a stable pH gradient & assist molecules (proteins) in migration.
A
Ampholyte
2
Q
Rate of migration of a charged solute in an electric field
A
Electrophoretic mobility
3
Q
Preferential movement of water in 1 direction through electrophoresis medium due to selective binding of 1 type of charge on the surface of the medium
A
Endosmosis
4
Q
Consist of sharply separated zone of
macromolecules
A
Electrophoretogram
5
Q
The result of zone electrophoresis
A
Electrophoretogram
6
Q
Migration of small charge ions
A
Iontophoresis
7
Q
Migration of charged macromolecules in a supporting medium
A
Zone electrophoresis
8
Q
The most prevalent electrophoretic technique used today
A
Zone electrophoresis
9
Q
Different types of electrophoresis
A
Paper electrophoresis
Capillary electrophoresis
Cellulose electrophoresis
Gel electrophoresis
10
Q
Particles are charged & are moving in an electrical field toward an electrode
A
Electrophoresis
11
Q
Usually used for Protein Determination
A
Electrophoresis
12
Q
Allows separation of substances in sample
A
Electric current
13
Q
Negatively charged (-) electrode
A
Cathode
14
Q
Positively charged (+) electrode
A
Anode
15
Q
Negatively charged (-) ion
A
Anion
16
Q
Positively charged (+) ion
A
Cation
17
Q
Cations are attracted to
A
Cathode
18
Q
Anions are attracted to
A
Anode
19
Q
Supplies negative charge (Cathode)
A
Black wire
20
Q
Supplies positive charge (Anode)
A
Red wire
21
Q
Provides the matrix for substances to be separated from
A
Supporting medium
22
Q
Contains the buffer/electrolyte solution
A
Subcell
23
Q
Both electrodes will be submerged in here
A
Buffer solution
24
Q
1st step of electrophoresis
A
Separation
25
Begins the moment it is subjected to the densitometer
Quantification
26
Add stains = can visualize
Detection
27
Factors affecting rate of migration (5)
1. Net electric charge of molecule
2. Size & shape of the molecule
3. Strength of electrical field
4. Properties of supporting medium
5. Temperature of operation
28
Produced when current flows through a medium that has resistance.
Heat
29
Produced heat results in ____ in theral agitation of the dissolved solutes and leading to a ___ in resistance and an ___ in current
Increase thermal agitation
Decreased resistance
Increase current
30
Supplies the constant current
Driving force (electrical power)
31
Separation of molecules in electrophoresis often requires high voltages
50-200 VDC
32
Common supporting medium
Cellulose acetate
Agarose gel
Polyacrylamide gel
33
Dry, brittle film composed of about 80% air space
Cellulose acetate
34
Homogenous medium w/ uniform pore size
and does not absorb the protein; can be used for serum protein electrophoresis
Cellulose acetate
35
most common analyte tested for electrophoresis in the PH
Protein
36
● Can also be stored for long periods
● Can only observe 5 fractions
Cellulose acetate
37
Widely used supporting medium for electrophoresis.
Agarose gel
38
Used as a culture medium for bacteria & electrophoresis
Agar
39
Does not produce electroendosmosis since it is neutral
Agarose gel
40
Advantages:
Good background for staining glycoprotein
Sharp bands
Offers good resolution
Cellulose gel
41
Referred to as page
Polyacrylamide gel
42
Separates proteins based on its charge &
molecular size
Polyacrylamide gel
43
Best supporting medium for serum
protein electrophoresis
Polyacrylamide gel
44
Separates proteins on the basis of surface charge & molecular size; not widely used
Starch
45
Polysaccharide extracted from seaweed
Agarose
46
Cross-linked polymer of acrylamide and cross-linkers such as bisacrylamide.
Polyacrylamide gel
47
Gel casted vertically
Polyacrylamide gel
48
Gel casted horizontally
Agarose
49
Potent neuro-toxic = best for testing; separate small molecules
Polyacrylamide gel
50
Used for DNA or protein separations
Polyacrylamide
51
Non-toxic = best for safety
Separate large molecules
Agarose
52
Commonly used for DNA separations. For the purpose of diagnosing genetic diseases, detecting bacteria and viruses, determining paternity, etc.
Agarose
53
Staining can be done AFTER pouring.
Polyacrylamide gel
54
Staining can be done BEFORE pouring the gel.
Agarose
55
Carries applied electrical current & set the pH of the electrophoresis
Buffer
56
Molecule whose net charge can be either positive/negative
Help establish a stable pH gradient and
assist molecules in migration
Ampholyte
57
Properties affecting ampholyte
pH and Ionic strength
58
pH at w/c a substance carries no net electric charge
Isoelectric point (pl)
59
Becomes positively charged and migrates toward the cathode
Binds more hydrogen ions
Buffer More Acidic Than pI
60
Becomes negatively charged and migrates toward the anode
Loses hydrogen ions
Buffer More Basic Than pI
61
Measure of the concentration of ions in the solution
Ionic strength
62
Higher Ionic Strength/Concentration
Bigger ionic cloud = slower mobility
63
Lower Ionic Strength/Concentration →
More current carried by particles = faster mobility
64
● serum protein separation
● Poor resolution, weak buffer
Barbitone buffer (around 8.0 pH)
65
● Enzyme separation
● Low buffering capacity - high conductivity
Phosphate buffer (around 7.0 pH)
66
● Nucleic acid separation
● Good resolution, high
buffering capacity, low conductivity
Tris-borate-EDTA buffer (TBE) - (pH around 8.0)
Tris-acetate-EDTA buffer (TAE) - (pH around 8.0)
67
● Protein separation
● High buffering capacity,
lox conductivity
Tris-glycerine buffer - (pH more than 8.0)
68
Degree of separation of substances
Resolution
69
Ability of buffer to resist pH changes
Buffer capacity
70
Ability of buffer to conduct electricity
Conductivity
71
Visualizes the macromolecules of interest under the UV light
Uses stains and densitometer for quantification
Detecting system
72
help in the visualization of the bands & macromolecules (proteins) that have migrated and the locations of the separated particles
Stains
73
Stains used to visualize proteins
● Amido Black
● Coomassie Brilliant blue
● Bromophenol Blue
74
Stains used to visualize DNA
● Ethidium Bromide
● Sybr Green
● Sybr Gold
75
Stains used to visualize lipoproteins
Sudan black
76
Stains used to visualize hemoglobin
● Ponceau Red
● Amido Black
● Coomassie Brilliant Blue
77
It measures the absorbance of concentration of the dye (stain) & protein fraction
Densitometry
78
Has the most negative charge & will migrate farthest towards the anode
Albumin
79
Occurs due to light passing from 1 medium to another medium
Refraction
80
Ability of the substance to bend
light
Refractivity
81
Ratio of the 2 speeds of light
Refractive index
82
The comparison of the velocity of light in air & the velocity of light in the solution.
Refractive index
83
Involves the measurement of electrical signals
associated w/ a chemical system that are incorporated into an electrochemical cell
Electrochemistry
84
Converts chemical energy to electrical energy through redox reaction
GALVANIC/ELECTROCHEMICAL CELL
85
Spontaneous & is responsible for the production of electrical energy
Redox reaction
86
A solution w/ a submerged electrode
Half cell
87
Allows electrical contact between the two solutions
Liquid junction/ salt bridge
88
Its purpose isn't to move electrons from the electrolyte, but to maintain charge balance
Liquid junction
89
Measure potential (voltage) under zero current
Potentiometry
90
Electrochemical titration in w/c the titrant is electrochemically generated & the endpoints is detected by amperometry = constant current
Coulometry
91
Measure current
Amperometry
92
Measure current after applying a potential
Voltammetry
93
Describes the electromotive force generated because of the hydrogen ion (H+)
at the glass tip
Nernst equation
94
Produce constant potential
Reference electrode
95
Produce varied potential
Indicator potential
96
Mercury surrounded by mercurous chloride submerged in a saturated potassium chloride solution
Calomel electrode
97
Silver electrode immersed in potassium chloride solution saturated w/ silver chloride solution
Silver or silver chloride
98
Best reference electrode for potentiometry
Silver/ silver chloride
99
Indicator electrode that can respond to individual types of anions or cations
Ion-selective electrode
100
Depends on the membrane or barrier composition used
Ionic selectivity
101
No sample dilution
Direct ISE
102
Has sample dilution
Direct ISE
103
Composed of both indicator &
reference electrodes
Combination Electrodes (pH & pCO2
Electrodes)
104
Selective for the detection of hydrogen
ions
PH electrode
105
pH electrode w/in a plastic jacket
pCO2 ELECTRODE
106
AKA Severinghaus Electrode or
Severinghaus-Stow Electrode
PCo2 electrode
107
The measurement of current after w/c a potential is applied to an electrochemical cell
Voltammetry
108
For Lead & Iron Testing
Anodic Stripping Voltammetry
109
3electrodes under voltametry
Working electrode
Reference electrode
Auxiliary electrode
110
Makes contact w/ analyte/sample = acts as indicator electrode
Working electrode
111
A half cell w/ a known reduction potential
Reference electrode
112
Sustains electrolysis = chemical decomposition using electricity
Auxiliary electrode
113
Facilitate the transfer of charge to and from the analyte
Working electrode
114
Involves the application of a constant current to generate a titrating agent
Coulometry
115
The time required to titrate a sample at a constant current is measured and is related to the amount of analyte in a sample by
Faraday’s equation
116
Measurement of the current flow produced by an oxidation-reduction reaction at a single applied potential
Amperometry
117
A measure of the cell current when the potential difference b/w indicator and reference electrodes is controlled
Amperometry
118
Measurement of osmolality of an aqueous solution (serum, plasma, and urine)
Osmometry
119
Measurement of the concentration of dissolved solute particles in a solution or fluid.
Osmometry
120
colligative properties related to osmometry
Vapor pressure
Boiling point
Osmotic pressure
Freezing poit
121
Increased boiling point = __ osmolality
Increased
122
Decrease vapor pressure = __ osmolality
Increase
123
Freezing point decrease =___ osmolality
Increase
124
Pressure that is acted on the solution to retain its volume
Osmotic pressure
125
Increased osmotic pressure = ___ molality
Increase
126
Directly proportional to osmolality
Osmotic pressure
Boiling point
127
Indirectly proportional to osmolality
Freezing point
Vapor pressure
128
Measurement is based on the change in electrical
resistance across an aperture when a particle in
conductive liquid passes through this aperture
Electrical impedance
129
Blood sample is divided into
Mix w diluent
Mix w diluent and cytochemical stains
130
Cell bath = Erythrocyte
(RBC) Count and Platelet Count
Mix w diluent
131
Lysed the RBCs only = Leukocyte (WBC) Count
Mix w diluent and cytochemical stains
132
proposed the idea of electrical impedance.
Wallace Coulter
133
Detects and measures changes in
electrical impedance or resistance produced by a blood cell (RBC, platelets, WBC), as it passes through an aperture or an electrical field
Coulter machines
134
Uses technology or principle that counts & size particles by measuring impedance/resistance
Beckman-Coulter Chemistry Analyzer
