Analytical Techniques and Automation Flashcards
1
Q
Describes the measurement principles used in the clinical chemistry laboratory
A
Analytic Techniques
2
Q
Examples of analytic techniques
A
Spectrophotometry
Electrochemistry
Electrophoresis
Chromatography
3
Q
Analytic techniques often used to determine concentrations of analytes in the CC lab
A
Spectrophotometry
Electrochemistry
4
Q
Described as photons of energy traveling in waves
A
Electromagnetic radiation
5
Q
Most recognizable forms of electromagnetic radiation
A
Light and radiant energy
6
Q
Other types of electromagnetic radiation
A
Gamma rays
X-rays
Microwaves
Ultraviolet radiation
Radiofrequency
Radiation
7
Q
The linear distance between any two equivalent points on a successive wave
A
Wavelength
8
Q
Unit used in the visible spectrum
A
Nanometer nm
9
Q
The relationship between wavelength and energy (E) is described by
A
Planck’s formula
10
Q
Planck’s formula
A
𝐸 = h𝑣
11
Q
Planck’s constant value
A
6.62 x10^-27 erg/sec
12
Q
Unit of energy in the centimeter gram second system unit
A
erg
13
Q
Is erg an SI unit?
A
No
14
Q
h in the Planck’s formula
A
Planck’s constant
15
Q
v in Planck’s in formula
A
Frequency
16
Q
Number of oscillations of the waveform in a second
A
Frequency
17
Q
Changes that occur in a period of time. Movement of waveform
A
Oscillation
18
Q
Relationship between wavelength and frequency
A
Inversely proportional
19
Q
Relationship between energy of electromagnetic radiation and wavelength
A
Inversely proportional
20
Q
Wavelength of visible region
A
400-700nm
21
Q
Wavelength of ultraviolet region
A
<400nm
22
Q
Wavelength of infrared region
A
> 700nm
23
Q
Visible region falls in between
A
Color violet (set at 400nm) and red (at 700nm)
24
Q
States that the concentration of a substance is directly proportional to the amount of light absorbed or inversely proportional to the algorithm of the transmitted light
A
Beer’s Law
25
Converts the radiant energy into equivalent electrical energy
Photodetector
26
Function of sample cuvette
Contain the sample
27
Function of light source
Strikes the sample in the cuvette
28
Formula for % transmittance
% transmittance = T/I x 100
29
T in % transmittance formula
Radiant energy transmitted/ transmitted light
30
I in % transmittance formula
Radiant energy incident of the sample/ incident light
31
Amount of energy absorbed by the sample
Incident light (I)
32
Radiant energy that strikes photodetector
Transmitted light (T)
33
It refers to the amount of light absorbed
Absorbance
34
Formula for absorbance
𝐴 = 𝜀 ×𝑏 × 𝑐
35
𝜀 in absorbance formula
Molar absorptivity
36
b in absorbance formula
Length of the light path through the solution
37
c in absorbance formula
Concentration of absorbing molecules
38
Relationship between absorbance and concentration of the absorbing molecules or analyte of interest
Directly proportional
39
Equipment used to measure the light transmitted by a solution
Spectrophotometer
40
Function of spectrophotometer
Determine the concentration of the light absorbing substance in the solution
41
Components pf spectrophotometer
light source
Monochromator
Sample cell or cuvet
Photodetector
Meter or read-out device
42
Function of light source
Provides polychromatic light
43
What is polychromatic light
Light of several wavelengths. Variation in the color formation
44
Light source for visible and near-infrared region use
Incandescent tungsten or tungsten-iodide lamp
45
Light source for UV region use
Dueterium lamp and memory arc lamp
46
Light source will depend on the wavelength. True or False?
True
47
2 types of light source
Continuum
Line
48
Type of light source that emits radiation that changes in intensity
Continuum
49
Examples of continuum light source
Tungsten (visible region)
Dueterium (UV region)
Xenon (visible and UV region)
50
Type of light source that emits a few discrete lines or bands of radiation
Line
51
Examples of line light source
Mercury and sodium vapor
lamps (UV and visible)
Hollow cathode lamp (atomic absorption spectroscopy/ spectrophotmetry)
52
Function of monochromator
Isolates individual wavelengths of light
53
When light source strikes the monochromator, light source will produce ______
Polychromatic lights
54
Wavelength in nanometers (nm) at peak transmittance
Nominal wavelength
55
Range of wavelength about one half peak transmittance
Spectral bandwidth (FWHM or full width at half peak maximum)
56
Area from one point of a wave to another
Bandpass
57
Total length of wavelength
Bandpass
58
Shape and material for sample cuvette
Round or square; made of material that is transparent to radiation
59
Path length of sample cuvette
1 cm
60
Types of materials used for cuvette and their respective region of use
Fused silica or quarts: UV region
Alumina-silicate glass: 350-2000 nm
wavelength
Plastic cuvette: visible region
61
Double-beam spectrophotometers has how many cuvette?
Two cuvets (one for the sample and one for the solvent)
62
Function of photodetector
Converts the transmitted radiant energy into an equivalent amount of electrical energy
63
Types of photodetector
Barrier-layer cell or photocell (selenide cell)
Phototube
Photomultiplier tube (PMT)
Phototransistors and Photodiode
64
The least expensive; temperature sensitive photodetector
Barrier-layer cell or photocell
65
Composition of Barrier-layer cell or photocell
Selenium on a plate of iron
66
Barrier-layer cell or photocell is mainly used in _____
Filter photometers
67
Photodetector that contains cathode and anode enclosed in a glass tube
Phototube
68
Photodetector that has photosensitive material that gives off electrons when light energy strikes it
Phototube
69
Most common type of photodetector
Photomultiplier tube (PMT)
70
Characteristics of Photomultiplier tube (PMT)
200 times more sensitive than the phototube
Highly sensitive to UV and visible radiation
71
Characteristics of Photodiode
Not as sensitive as PM tube but has excellent linearity and speed
72
Beam of light that strikes the photodetector reflect the amount of analyte present in the sample
Linearity
73
Concentration of analyte can immediately be displayed in the digital meter because it can immediately convert amount of radiant energy into an equivalent energy
Speed
74
Function of meter or read-out device
Displays output of the detection system
75
Examples of meter or read-out device
Digital meters
d’Arsonval meters
Recorders
Light-emitting diodes (LEDs)
Cathode-ray tubes (CRTs)
Liquid crystal displays (LCDs)
76
Simplest type of absorption spectrometers
Single-beam spectrophotometer
77
Function of single-beam spectrophotometer
Measure one measurement at a time at one specified wavelength
78
Components of single-beam spectrophotometer
Light source
Monochromator
Sample cuvette
Photodetector (PM tube)
Read out device or meter
79
Components of double-beam spectrophotometer
Light source
Monochromator
Sample cuvette
Reference Cuvette
Photodetector (PM tube)
Read out device or meter
80
Spectrophotometer design that uses 2 photodetectors, 2 sample cuvettes
Double beam in space
81
Spectrophotometer design that uses 1 photodetector; chopper is used to pass the monochromatic radiation through the sample cuvette and then to the reference cuvette
Double-beam in time
82
A device that rotates or breaks up radiation beams so the beam of light can pass through the photodetector
Chopper
83
This implies that a photometer is measuring at the wavelength that it is set to
Wavelength or photometric accuracy
84
Material used to measure wavelength or photometric accuracy
Special glass-type optical filters:
Didymium glass (600 nm)
Holmium oxide (360 nm)
85
A test using glass filters or solutions that have known absorbance values for a specific wavelength
Absorbance check
86
The ability of a photometric system to yield a linear relationship between the radiant power incident upon its detector and the concentration
Linearity
87
Linearity is monitored using _____
Optical filters or solutions
88
Any light that impinges upon the detector that does not originate from a polychromatic light source (an interference)
Stray light
89
The presence of stray light is checked using _____
Special cut-off filters
90
Measures concentration by detecting the absorption of electromagnetic radiation by atoms rather than by molecule
Atomic Absorption Spectrophotometer
91
When using AAS, sample should be ___
Atomized
92
Light source for AAS
Hollow-cathode lamp
Electrodeless discharge lamp (new edition)
93
Function of chopper in AAS
Modulate the light beam that would strike the sample cell that comes in flame
94
Photodetector for AAS
Photomultiplier tube
95
Application of AAS
To measure concentration of trace metals ( lead, mercury, cadmium)
96
Components of single-beam AAS
Light Source
Chopper
Sample
Monochromator
PM tube (Photomultiplier tube)
Readout/ Meter
97
AAS that does not require a burner to produce a flame but uses electric current
Flameless AAS
98
Method used to measure light emitted by excited atoms
Ion Selective Electrode
99
Application of ion selective electrode
To measure concentrations of sodium, potassium, and lithium
100
Measures the concentrations of solutions that contains fluorescing molecules
Fluorometry
101
Light source for fluorometry
Mercury (for filter fluorometers)
Xenon arc (for spectrofluorometers)
102
Fluorometer that use filter as monochromator (primary and secondary filter)
Filter fluorometers
103
Fluorometer that use prisms or gratings
Spectrofluorometers
104
Photodetector for fluorometry
Photomultiplier tube
105
Basic component of Filter fluorometer
Source
Attenuator
Primary filter
Sample holder
Secondary filter
Detector (PM)
Readout
106
Filters used in filter fluorometer and their function
Primary filter: selects wavelength of light that is best absorbed by sample
Secondary filter: passes the longer wavelength of light to photodetector
107
Advantages of fluorometry
Specific and sensitive
108
Disadvantage of fluorometry
Sensitive to environmental changes resulting to Quenching (decrease in fluorescence due to these changes)
109
Environmental changes causing quenching
Use of contaminated chemicals
Change in solvents
Use of UV light which causes photochemical changes
Change in temperature
110
The emission of light as a result of chemical reaction
Chemiluminescence
111
How does chemiluminescence differ from fluorescence?
No excitation radiation is required and no monochromators are needed
112
It is the production of electromagnetic radiation when a chemical reaction yields an excited product
Chemiluminescence
113
Advantages of chemiluminescence
Subpicomolar detection limits, speed, ease of use, and simple instrumentation
114
Disadvantage of chemiluminescence
Impurities can degrade sensitivity and specificity
115
Principle of turbidimetry and nephelometry
Based on the scattering of radiation by particles in suspension
116
Applications of turbidimetry and nephelometry
Measurement of antigen-antibody reactions, prealbumin, and other serum proteins
117
It is the measurement of the light scattered by a particulate solution
Nephelometry
118
3 types of light scatters
Rayleigh theory
Mie theory
Rayleigh-Debye theory
119
This theory states that when the wavelength of light is greater than the diameter of the particle, then there is symmetrical distribution of light scattering
Rayleigh theory
120
According to rayleigh theory, the minimum light scatter occurs at how many degrees to the incident light?
90 degrees
121
This theory states that if the wavelength of light < the particle diameter (d > 0.1 λ), then the light scatters forward
Mie theory
122
This theory states that if the wavelength of light is approximately the same as the particle size, more light scatters in the forward direction than in other direction
Rayleigh-Debye theory
123
Device used to measure a concentration of a solution using the analytical technique nephelometry
Nephelometer
124
Components of nephelometer
Light source
Collimator
Monochromator
Sample cuvette
Photodetector
125
Light source for nephelometer
Mercury-arc lamp
Tungsten- filament lamp
Light-emitting diode
Laser
126
Function of collimator
Narrows or control a beam of light
127
Determines the amount of light blocked by a suspension of particles
Turbidimetry
128
Applications of turbidimetry
It is used in microbiology analyzers, coagulation analyzers, and is used to quantify protein concentration in biologic fluids such as urine and CSF
129
What is the difference between nephelometry and turbidimetry?
Nephelometry detects (right-angle or forward) scattered light, and turbidimetry measures a reduction of light transmitted in the forward direction
130
Electrochemistry techniques
Potentiometry
Coulometry
Amperometry
Voltammetry
131
Involves the measurement of the current or voltage generated by the activity of specific ions
Electrochemistry
132
Most widely used technique in clinical measurements
Potentiometry
133
Measurement of potential (voltage) between two electrodes in a solution
Potentiometry
134
The two electrodes in potentiometry
Reference Electrode
Indicator Electrode
135
Electrode with a constant voltage
Reference Electrode
136
Most used reference electrode in the laboratory
Calomel and Silver/Silver chloride
137
Measuring electrode
Indicator Electrode/Analytical Electrode
138
Can be calculated from the measured potential difference between 2 electrodes
Concentration of Ions
139
Replaced the Flame Photometry
Ion-Selective Electrode
140
Membrane used to measure sodium
Glass aluminum silicate
141
Membrane used to measure potassium
Valinomycin Gel
142
Membrane used to measure calcium and lithium
Organic Liquid Ion Exchangers
143
Membrane used to measure carbon dioxide and ammonia
Gas Electrodes
144
Membrane used to measure urease and glucose oxidase
Enzyme Electrodes
145
Two type of ISE
Direct ISE
Indirect ISE
146
ISE that does not require sample dilution
Direct ISE
147
ISE that requires sample dilution before the analysis phase
Indirect ISE
148
Used to measure hydrogen ion (concentration) activity
pH electrode
149
Internal Reference Electrode
Silver/silver chloride
150
External Reference Electrode
Calomel electrode
151
A pH electrode within a plastic jacket (has sodium bicarbonate buffer and gas-permeable membrane)
pCO2
152
Measures the quantity of electricity (in coulombs) needed to convert an analyte to a different oxidation state
Coulometry
153
Applications of coulometry
To measure chloride ion in serum, plasma, CSF, and sweat samples
154
It is the measurement of the current flow produced by an oxidation-reduction reaction
Amperometry
155
Applications of amperometry
To measure chloride ion in serum, plasma, CSF, and sweat samples; pO2 electrode blood gas analyzers
156
Gas sensing electrode
pO2 Electrode
157
Application of pO2 electrode
To measure the partial pressure of oxygen in the blood
158
it is a method in which a potential (voltage) is applied to an electrochemical cell and the resulting current is measured
Voltammetry
159
Used to measure heavy metals such as lead
Anodic Stripping Voltammetry
160
Measurement of the number of dissolved particles in a fluid
Osmometry
161
Osmotically active particles
Glucose
Urea Nitrogen/ Blood Urea Nitrogen (BUN)
Sodium
162
Effects of increased osmolality
Osmotic pressure increases
Boiling point increases
Freezing point decreases
Vapor pressure decreases
163
Is used to measure the concentration of solute particles in a solution
Osmometer
164
The process of separating the charged constituents of a sample by means of an electric current
Electrophoresis
165
Electrophoresis is the separation of charged compounds based on their
Electrical charge
166
A substance that can either have a negative, zero or positive charge depending on the conditions
Amphotheric
167
Compounds that when dissolved in water can act either as acid or as a base
Ampholytes
168
Negatively charged ions
Anion
169
Positively charged ions
Cation
170
Ions that are neutral and have both positive and negative charges at different locations throughout the molecule
Zwitterions
171
Negatively charged electrode
Cathode
172
Positively charged electrode
Anode
173
Factors affecting the mobility of particles
Net charge of the particle
Size and shape of the particle
Strength of the electric field
Chemical and physical properties of the medium
Electrophoretic temperature
174
Migration of small ions
Iontophoresis
175
Migration of charged macromolecules in a porous support medium
Zone Electrophoresis
176
Components of electrophoresis
Power supply
Buffer
Support Medium
Sample
Detecting System
177
Function of power supply
Supplies constant current or voltage in the system
178
Known as the driving force in electrophoresis
Power supply
179
Why does the power supply named as the driving force in electrophoresis?
Because this drives the molecules through the support medium
180
Function of buffer
Provide ions that will enable the movement of current and migration of particles
Maintain the pH at a relatively constant value
181
A mixture of proton-donating and proton-accepting substances that functions to maintain the pH at a constant value
Buffer
182
Barbital (veronal) pH
8.6
183
Tris-boric EDTA pH
8.7
184
Cation migrates to the
Cathode
185
Anion migrates to the
Anode
186
pH will influence the charge of the analyte. True or False?
True
187
Relationship between ionic strength and mobility
LOW I.S = more charge will be carried = faster mobility
HIGH I.S = less charge will be carried = slower mobility
188
A network of interacting fibres or a polymer that is solid but traps large amount of solvent in its pores or channels inside
Support media
189
The support media must not interact with the analyte, it is just supposed to support the analyte to pass through it. True or False?
True
190
Examples of support media
Cellulose Acetate
Agarose Gel
Polyacrilamide Gel
191
Cellulose that is acetylated from cellulose acetate by treating it with acetic anhydride
Cellulose acetate
192
Cellulose acetate separates serum proteins into how many bands
5 bands
193
5 bands produced when using cellulose acetate
Albumin
Alpha-1
Alpha-2
Beta
Gamma
194
The support media of choice
Cellulose acetate
195
Useful in doing electrophoresis with proteins that can be separated with 5 bands
Isoelectric Focusing
196
Used as a purified fraction of agar that comes from red algae
Agarose gel
197
Unique features of agarose gel
It is neutral and does not produce electroendosmosis
198
Agarose gel separates proteins into how many bands?
10-15 bands
199
Support medium used to separate protein based on charge and molecular size
Polyacrilamide gel
200
Polyacrilamide gel separates serum proteins into how many bands?
20 or more bands
201
Result of electrophoresis consisting of separated strands of a macromolecule
Electrophoretogram
202
Detecting system that can already be conducted if the samples are already dyed or stained
Direct observation
203
The simplest way of detection in electrophoretic system
UV visualization
204
A device that measures the degree of darkness of a photographic or semitransparent material or of a reflecting surface
Densitometer
205
Examples of electrophoretic detecting system
Electrophoretogram
Direct observation
Staining
Radioactive dye
UV visualization
Densitometer
206
Applications of electrophoresis
DNA Fractionation
Isoenzyme Determination
Protein Fractionation
207
Process of forming electric cloud that prevents analytes or particles from migrating into the support medium
Electroendosmosis
208
Separation is determined through the speed of the analyte’s migration
Isoenzyme determination
209
Separate complex mixture on basis of different physical interactions between individual compound and stationary phase of the system
Chromatography
210
Basic components of chromatography and their functions
Mobile Phase: carries complex mixture
Stationary Phase: through which mobile phase flows
Column: holds the stationary phase
Eluate: separated component
211
Modes of separation in chromatography
Adsorption
Partition
Steric Exclusion
Ion exchange
212
2 classifications of chromatography based on stationary phase
Planar chromatography
Column chromatography
213
Classification of chromatography where the stationary phase is coated with a sheet of paper or bound to glass or plastic plate
Planar chromatography
214
Classification of chromatography where the stationary phase is packed into tube or coated onto the inner surface of the tube/column
Column chromatography
215
Examples of Planar chromatography
Paper chromatography
216
Examples of Column chromatography
Thin layer chromatography
Gas chromatography
Liquid chromatography
217
Application of Paper chromatography
Fractionation of sugar and amino acid
218
Application of Thin-Layer chromatography
Drug screening
219
Application of Gas chromatography
Separate mixture of compounds that are volatile made or can be made volatile
220
Application of Liquid chromatography
Uses pressure for fast separation of thermolabile substance
221
Why do we need to separate thermolabile substances immediately?
Thermolabile substances must be separated immediately, because when it is exposed to high temp. they will be destroyed and goes unstable that can cause us not to recover anything
222
How can we differentiate a gas chromatography to a liquid chromatography?
We can differentiate them based on their mobile phase
223
How can we differentiate planar and column chromatography?
We can differentiate them based on their stationary phase
224
Forces the mobile phase through the column
Pumps
225
Holds the stationary phase
Columns
226
Introduce the sample into the mobile phase
Sample injectors
227
Produce an electronic signal proportional to the concentration of separated component
Detectors
228
Most common photodetector used in spectrophotometer
Photomultiplier tube
229
Save the measurement of elutions
Recorders
230
Elution strength of the mobile phase is constant
Isocratic elution
231
2 distinct portion of mass spectrophotometer
Fragmentation
Ionization
232
Separate the components of a mixture
Fragmentation
233
Methods use to ionize samples
Electron Spray Ionization (ESI)
Matrix Assisted Laser Desorption Ionization (MALDI)
234
Analyzers used to measure mass-to-charge ratio
Quadrupole mass analyzers
Iron trap analyzers
Time of flight analyzer
