Deck 9: Optical Techniques

Question 1

Which one of the following components is found only in a fluorometer used to make measurements of emitted fluorescent light as opposed to in a spectrophotometer used to make measurements of absorbed/transmitted light?

A) Monochromator
B) Cuvets as sample holders
C) An excitation source
D) Detector

Answer 1

An excitation source

Question 2

A fluorometric measurement technique used to detect immunologic reactions that measures fluorescence produced by the rotation of small versus large fluorescent-labeled molecules and their relation to analyte concentration in an unknown sample is known as:

A) phosphorimetry.
B) reflectance photometry.
C) nephelometry.
D) fluorescence polarization.

Answer 2

fluorescence polarization.

Question 3

Which one of the following wavelengths is within the "near" ultraviolet region of the electromagnetic spectrum?

A) 320 nm
B) 450 nm
C) 540 nm
D) 690 nm

Answer 3

320 nm

Question 4

A blood specimen is obtained from an individual who is known to have elevated lipids and the serum appears milky.This individual's physician has ordered an estrogen immunoassay that your laboratory performs using a nephelometric technique.Which one of the following statements is correct concerning the possible outcome of this assay?

A) Certain components of this individual's sample will likely produce excess fluorescence by the phenomenon called "solvent effect" in the nephelometric assay.
B) The lipemic specimen will produce interfering background light intensity and excess light scatter in this type of assay.
C) The sample flow rate will be affected through the analyzer's nebulizer because of the altered density of the sample.
D) Elevated serum lipids will not interfere with this assay because nephelometry is not affected by any type of sample variability.

Question 5

Which one of the following statements best describes Beer law?

A) The concentration of a substance is inversely proportional to the logarithm of transmitted light.
B) Absorbance is inversely proportional to the logarithm of the concentration.
C) Transmitted light is directly proportional to the concentration of a substance in solution.
D) Intensity of incident light divided by intensity of transmitted light equals concentration.

Question 6

In spectrophotometric determinations,which of the following is the formula for calculating the absorbance of a solution?

A) (Molar absorptivity * length of light path)/concentration in mol/L
B) (Molar absorptivity * concentration in g/dL)/length of light path
C) Molar absorptivity * length of light path * concentration in mol/L
D) (Length of light path * concentration in g/dL)/molar absorptivity

Question 7

Which component of a generic spectrophotometer isolates radiant energy of a specific wavelength (spectral isolation)and excludes that of other wavelengths?

A) Monochromator
B) Entrance slit
C) Cuvet
D) Light source

Question 8

In regard to absorption photometry,absorbance is calculated from transmittance by which one of the following formulae?

A) A = log %T - 2
B) A = log T
C) A = -log T
D) A = abc

Question 9

In regard to atomic absorption (AA)spectrophotometry,which one of the following statements is correct?

A) The technique combines laser-induced fluorometry and particle light scattering analysis to differentiate molecules,cells,or particles by size and shape.
B) The technique measures concentration through the detection of absorbance of electromagnetic radiation by atoms of elements instead of molecules.
C) Oxidation of an organic compound such as luminol induces an excited state,and light is emitted when the electron returns to the ground state.
D) The amount of light scattered at right angles to the incident light is directly proportional to the concentration of the analyte of interest.

Question 10

Atomic absorption spectrophotometry is an emission technique that is used frequently in clinical laboratories to measure:

A) complex antigen-antibody reactions.
B) rotational relaxation of bound fluorophores.
C) elements.
D) high-molecular-weight compounds.

Question 11

A standard calibrator solution of glucose has a concentration of 125 mg/dL and gives an absorbance of 0.20.Assuming a linear reaction,what is the concentration of glucose in a patient's serum that has an absorbance of 0.24?

A) 104 mg/dL
B) 138 mg/dL
C) 150 mg/dL
D) 180 mg/dL

Question 12

A molecule is said to fluoresce when it absorbs light at one wavelength and emits light of:

A) a lower energy.
B) a shorter wavelength.
C) a higher energy.
D) the same wavelength.

Question 13

Which one of the following statements concerning nephelometry is correct?

A) Nephelometry is the measure of the concentration of particles by measuring the amount of incident light blocked by the particles.
B) In a nephelometric measurement,light blocked by solutes in the solution at 180° from the incident light is measured by a photodetector.
C) The decrease in the intensity of scattered light is directly proportional to the number of particles in the solution,and the measurement of this decrease is called nephelometry.
D) In a nephelometric procedure,the measurement of scattered light that is not in the direct path of the transmitted light is made at right angles to the incident light.

Question 14

A manual stat protein analysis is performed on a patient sample.The patient's unknown sample had an absorbance reading of 0.6.The protein calibrator with an absorbance reading closest to the unknown's absorbance had a concentration of 6 g/dL and an absorbance reading of 0.5.Applying Beer law,calculate the unknown protein concentration in the patient sample.

A) 6 g/dL
B) 7.2 g/dL
C) 8 g/dL
D) 5.5 g/dL

Question 15

Which one of the following would be a limitation in a nephelometric measurement that might produce faulty values?

A) Excess antigen in the antigen/antibody reaction
B) Too much fluorophore in the analyte/fluorescing compound mixture
C) Incorrect filter used to isolate a specific wavelength
D) Absorption by a closely related atomic species

Question 16

The light source in an atomic absorption spectrophotometer is typically a:

A) prism.
B) tungsten light.
C) laser light.
D) hollow cathode tube.

Question 17

When selecting a photometric technique that is more sensitive to a low concentration of analyte,a laboratory director compares absorbance measuring devices against those that measure fluorescence intensity.Which technique is considered more sensitive in its measurements and why?

A) Absorbance measurements are more sensitive because of the ability of the monochromator to isolate very specific parts of the spectrum.
B) Fluorescence intensity measurements are more sensitive because the fluorophores used are very specific to the analytes they bind to and will bind to small analytes easily.
C) Fluorescence intensity measurements are more sensitive because of the use of intense light sources,signal filtering,and sensitive light emission photometers.
D) Absorbance measurements are more sensitive because the flame can break down complex compounds into elements that are specifically measured.

Question 18

Laboratory analysis of free thyroxine (fT₄)in your chemistry laboratory is performed with a conventional fluorometric analyzer,which uses a laser light as the excitation source.You have received a sample from an individual known to have a very elevated fT₄ level.For the sample's value to be within linear range of the assay,you dilute the sample by a factor of 10 and proceed with the analysis.Upon reviewing the results,you note that the value of the diluted sample is reported as "none detected." What is the likely reason for this result?

A) Hormones cannot be assayed using fluorometry because they will produce concentration quenching.
B) The dilution of the sample has lowered produced photobleaching by the intense light source and photodecomposition of the analyte.
C) Unwanted background fluorescence due to elevated protein hormone has produced a false-negative result.
D) High intensity light emission can lead to pulse pileup in photomultiplier tubes,leading to an underestimation of the true light emission intensity.

Question 19

A monochromator's filter must be tested to determine whether it can isolate a specific part of the electromagnetic spectrum.Percent transmittance (%T)of light is plotted against a specific wavelength in nanometers.It is noted at which wavelengths 50%T (one half of the peak transmittance)occurred.This type of test assesses the filter's:

A) linearity.
B) spectral bandwidth.
C) calibration.
D) absorbance spectrum.

Question 20

There are several interferences that can affect or limit fluorescence measurements.For example,the inner filter effect:

A) occurs when the solution absorbs greater than 2% of the exciting light,producing a nonlinear response between concentration and fluorescence emission.
B) involves fluorophores that have overlapping excitation and emission spectra and that are susceptible to loss of detection because of background light scatter.
C) involves solvents such as ethanol that cause appreciable fluorescence due to the interaction of the fluorophore with the solvent.
D) is caused by light scattering of proteins and other macromolecules in the sample matrix,which might cause unwanted background fluorescence.

Question 21

A light-beam chopper in a double-beam-in-time spectrophotometer is used to:

A) isolate light of a desired wavelength.
B) compensate for light source variation.
C) atomize a sample on a carbon rod in an enclosed chamber.
D) provide polarized light.

Question 22

The visible spectrum is composed of those wavelengths between 380 and 750 nm.

Question 23

Which two optical techniques are combined operationally in a flow cytometer?

A) Laser-induced fluorometry and light scattering techniques
B) Atomic absorption spectrophotometry and light scattering techniques
C) Spectrophotometry and fluorescence polarization techniques
D) Fluorescence polarization and laser-induced fluorometry

Question 24

In a fluorometric measurement,the difference between the maximum wavelength of the excitation light and the maximum wavelength of the emitted fluorescence light is a constant referred to as:

A) Beer law.
B) chemiluminescence.
C) polarization.
D) the Stokes shift.

Question 25

Which one of the following optical techniques is not considered an emission technique?

A) Phosphorimetry
B) Nephelometry
C) Luminometry
D) Fluorometry