Optical Techniques

Question 0

wavelength is measured in terms of _______

Answer 0

nanometers (nm)

Question 1

the distance traveled by one complete wave cycle

Answer 1

wavelength

Question 2

the number of cycles per second, inversely proportional to wavelength

Answer 2

frequency

Question 3

the peak height of a wave

Answer 3

amplitude

Question 4

wavelength that has a high frequency and high energy (400 nm)

Answer 4

short wavelength

Question 5

wavelength that has a low frequency and low energy (700 nm)

Answer 5

Long wavelength

Question 6

wavelength (in nm) of visible light

Answer 6

380-800 nm

Question 7

wavelength (in nm) of ultraviolet light

Answer 7

<380 nm

Question 8

The visible colors correspond to the wavelenghts that are _______ to our eyes

Answer 8

transmitted

Question 9

Only colors we can see are transmitted to the eyes, others are ______ but not seen. This color is the complementary color of the transmitted color.

Answer 9

absorbed

Question 10

these experiements were with the interaction of monochromatic (one wavelength) electromagnetic radiation with varying thickness of absorbing material

Answer 10

Lambert’s Experiments

Question 11

______ is inversely and logarithmically related to %T

Answer 11

absorbance

Question 12

experiments with transmittance of a solution under differing concentrations

Answer 12

Beer’s Experiments

Question 13

As concentration increases, absorbance increases linearly and transmittance decreases exponentially

Answer 13

Beer’s Experiments

Question 14

A=abC , state the law and what each letter represents

Answer 14

Beer’s Law; A=absorbance, a= molar absorptivity constant in g/L, b=path length of cuvette, C=concentration

Question 15

Beer’s Law is not valid if…(5 limitations)

Answer 15

1 very elevated concentrations are measured
2 incident light is not monochromatic
3 solvent absorption is significant compared to solute absorption (HIL)
4 radiant energy is transmitted by other mechanisms (stray light)
5 sides of the cuvette are not parallel

Question 16

function is to be a source of radiant energy

Answer 16

light source

Question 17

Four types of light sources

Answer 17

tungsten filament with halogen vapor, ultraviolet, hollow cathode lamp, and laser

Question 18

function is to focus light into a narrow beam

Answer 18

entrance slit

Question 19

function is to isolate the desired wavelength and exclude others

Answer 19

monochromator

Question 20

3 types of monochromators

Answer 20

glass filters, prisms, and diffraction gratings (used in lab)

Question 21

used for readings in the visible and near visible range of spectrum

Answer 21

colored agent of a glass filter

Question 22

augment wavelengths in phase, destroy those out of phase

Answer 22

interference filters (glass)

Question 23

eliminate transmission of wavelengths above or below a set wavelength

Answer 23

glass cut-off filters

Question 24

made of glass or quartz, these do not give a linear dispersion of light, creates bends in light

Answer 24

prisms

Question 25

a diamond point used to cut tiny furrows into aluminized face of perfect, flat piece of glass; precise angle and distance; will bounce off at different angles to determine which light will go through

Answer 25

diffraction grating

Question 26

function is to determine the bandpass (bandwidth) or range of wavelengths that will reach the sample, last line of defense before light hits the sample, defined as the bandwidth of the absorbance curve at a point equal to one-half of the max absorbance.

Answer 26

exit slit

Question 27

function is to act as a receptacle for the sample

Answer 27

cuvette

Question 28

______ cuvettes are used for visible measurements

Answer 28

glass

Question 29

_______ cuvettes are used for UV measurements

Answer 29

quartz

Question 30

function is to convert transmitted light energy to electrical energy

Answer 30

detector

Question 31

thin layer of semiconductor (selenium) deposited on metal base (iron), selenium coated with thin silver lacquer and covered by protective glass, encased in plastic

Answer 31

barrier layer cell (detector)

Question 32

semiconductors which change their charged voltage upon being struck by light; voltage change is converted to current (electron flow) and measured

Answer 32

Photodiode

Question 33

photomultiplier tube components

Answer 33

cathode, anode, photosensitive electrodes, encased in evacuated glass envelope

Question 34

Light energy strikes cathode, electrons are emitted and focus on first dynode; dynode emits an increased number of electrons, and each successive dynode does the same; final amplification may be 10^6 times more than initial energy

Answer 34

Photomultiplier Tube (PM)

Question 35

why are PM tubes preferred detectors for clinical laboratory spectrophotometers?

Answer 35

use a small amount of transmitted light to multiply the signal and expand the range of absorption vs concentration. This lessens the need to dilute a sample.

Question 36

function is to display the electrical signal from the detector in an understandable form; may be an LED, meter, recorder, printer, digital display, etc.

Answer 36

readout device

Question 37

3 types of spectrophotometers

Answer 37

single beam, double beam (in time/in space/dual), and recording

Question 38

Most common spectrophotometer

Answer 38

single beam

Question 39

advantage of double beam spectrophotometers

Answer 39

Useful when you need 100% accuracy

Question 40

These spectrophotometers run with a reference and are twice as big as a single

Answer 40

double beam and split (dual) beam (see picutres page 14 in notes)

Question 41

Quality control 1 purpose and 2 method for wavelength accuracy

Answer 41

1 to ensure wavelength is really going through the sample | 2 replacement of source lamp, holmium oxide (rare earth) glass filter, didymium filter

Question 42

Quality control 1 purpose and 2 method for band width

Answer 42

1 monitor range of wavelengths | 2 use mercury vapor lamp to verify

Question 43

Quality control 1 purpose and 2 method for stray light

Answer 43

1 no ambient room light | 2 use glass cut-off filters to verify

Question 44

Quality control 1 purpose and 2 method for linearity

Answer 44

1 make sure absorbance vs concentration | 2 didymium filter

Question 45

Quality control 1 purpose and 2 method for photometric accuracy

Answer 45

1 checking performance of instrument/spectrophotometer | 2 National Bureau of Standards (NBS) glass filters or potassium dichromate solutions

Question 46

failure to get reading in spect

Answer 46

usually electrical

Question 47

erratic readings in spec

Answer 47

improper warm-up

Question 48

incorrect wavelength in spec

Answer 48

we want 450 nm but can be a misaligned monochromator

Question 49

scattered light in spec

Answer 49

scratched or improper cuvette

Question 50

unsteady baseline in spec

Answer 50

grounding problem; when you zero instrument get crazy results

Question 51

dark current in spec

Answer 51

residual current when instrument is turned off

Question 52

emission of light by an atom or molecule after absorption of an excitation photon

Answer 52

fluorescence

Question 53

Emitted light has a _______ wavelength and _______ energy than the excitation light

Answer 53

longer, less

Question 54

______ detects emitted light and follows Beer's law

Answer 54

fluorometry

Question 55

Why do fluorometers have a right angle design?

Answer 55

To minimize the amount of excitation light that may reach the detector, improving sensitivity and specificity of fluorometers

Question 56

Technique where a chemical reaction (oxidation) of an organic compound causes immediate light flash

Answer 56

Chemiluminescence

Question 57

Measures the decrease in percent transmittance through a solution; increased conc of particles=less light transmitted through solution; follows Beer's Law and assays may be performed on a spectrophotometer

Answer 57

Turbidimetry

Question 58

Measures the amount of light that is scattered toward a detector at a variety of angles (not 0 or 180)

Answer 58

Nephelometer

Question 59

Two common chemiluminescent materials used

Answer 59

Luminol and acridinium ester (AE)