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abft exam > 6: Color Tests, Spectrophotometry, Immunoassays > Flashcards

6: Color Tests, Spectrophotometry, Immunoassays Flashcards

1
Q

Trinder’s Reagent for Salicylate

A

Tridner’s reagent: mercuric chloride, water, 1 M HCl, ferric nitrate

Purple = salicylate

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2
Q

FPN Test for Phenothiazines

A

Ferric Chloride, Perchloric Acid, Nitric Acid

Color results depend on specific phenothiazine (pink red, orange, violet, blue)

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3
Q

20 % NaOH + pyridine + heat

A

Red in pyridine layer = trichloro +

Used for chloral hydrate, chloroform

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4
Q

Conway Diffusion for Ethanol

A

Outer ring = sample
Inner ring = potassium chromate

Green = ethanol (or other reducing agent)

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5
Q

Paraquat/ Diquat

A

0.1% sodium dithionite in 1 M NaOH

Blue = paraquat
Green = diquat

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6
Q

Reinsch test for Heavy Metals

A

Copper wire + acidified and heated sample matrix result in black or silvery deposits/staining

Dull black = arsenic
purple black = antimony
shiny black = bismuth
silver =mercury

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7
Q

Marsh Test for Arsenic

A

Covert arsenic into arsine gas and deposit it into a black film

Black = arsenic

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8
Q

technique that incorporates the binding reaction of a target substance (antigen) w/ an antibody

A

Immunoassay

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9
Q

In immunoassays, the drug of interest

A

Antigen

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10
Q

In immunoassays, the part of the antigen that is recognized by the immune system

A

Epitope

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11
Q

In immunoassays, part of antibody that binds with antigen

A

Paratope

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12
Q

In immunoassays, antibodies are the same as

A

immunoglobulions (Ig)

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13
Q

These are produced by animals in antiserum.

A

Polyclonal antibodies

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14
Q

the concentration of an antibody as determined by finding the highest dilution at which it is still able to cause agglutination of the antigen

A

Titer

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15
Q

What is produced when an animal is given antigen (hooked with protein)

A

Monoclonal antibodies

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16
Q

Type on immunoassay that allows measurement of labeled antigen w/o separating bound and free antigen

A

Homogeneous

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17
Q

When two substances (antigen and labeled antigen) compete for the same antibody binding sites

A

Competitive Binding

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18
Q

Type on immunoassay that requires separation of bound and free antigen before labeled antigen is measured

A

heterogeneous

*more common; think ELISA

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19
Q

Homogenous Immunoassay technique

-When labeled drug is bound to antibody, enzyme is inactive
-When enzyme is active (unbound bc specimen had antigens that bound) NAD is converted to NADH (measured at 340 nm)

A

Enzyme Multiplied Immunoassay Technique (EMIT)

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20
Q

Homogenous Immunoassay technique

When Fluorescein-linked drug is bound to antibody, the fluorescein label does not rotate freely and the absorbed polarized light is emitted; when the label is free, it rotates in solution and the amount of emitted light is reduced

A

Fluorescence Polarization Immunoassay (FPIA)

*Think Fluorescein, polarized light = FP

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21
Q

Homogenous Immunoassay Technique

-Genetically engineered fragments of e-coli as enzyme label; activity of the enzyme requires an enzyme acceptor and an enzyme donor

-Reassociated enzyme hydrolyzes CPRG to CPR and galactose; CPRG does not absorb at 570 nm, but CPR does

-If a drug is found in the donor’s sample, the unbound Enzyme Donor part reassembles and reacts with the substrate and causes a change in the color absorbance.

A

Cloned Enzyme Donor Immunoassay (CEDIA)

*Think e-coli = C E

22
Q

Homogenous Immunoassay Technique

-Label = microparticle w/ several linked drugs
-No drugs present: microparticle binds multiple antibodies to form large aggregates (increase absorbance w/ time)

A

Kinetic Interaction of Microparticle in Solution (KIMS)

*Think: microparticle = aggregate; microparticle = M

23
Q

Heterogeneous Immunoassay Technique

Antibody coated wells; add analyte and labeled analyte, incubate, wash material, add substrate, develop color, add stop acid, measure color

A

Enzyme Linked Immunosorbent Assay (ELISA)

24
Q

Quants antigens between two layers of antibodies (capture and detection); antigen must have at least two epitopes capable of binding

A

Sandwich ELISA

25
Q

excessive antigen interferes with ability of detection antibodies to bind (resulting in reduced signal)

A

Hook Effect

26
Q

the degree of response in an immunoassay to a substance other than the analyte of interest

A

% cross reactivity

Cross-reactivity

(concentration reading of assay analyte)/(concentration of cross-reactivity analyte) x 100

27
Q

measures ability of test to correctly ID the positive cases

A

Sensitivity

*think opposite; positive = seNsitivity

Sensitivity (%)=(true positive)/((true positive+false negative)) x 100

28
Q

measures ability of test to correctly ID negative cases

A

Specificity

*think opposite; negative = sPecificity

Specificity (%)=(true negative)/((true negative+false positive)) x 100

29
Q

the study of absorption and emission of light by matter as a function of wavelength

A

Spectroscopy

30
Q

method to measure how much a chemical substance absorb light; done so by measuring the intensity of light as a beam passes thru a sample

A

Spectrophotometry

*think reading ELISA plates

31
Q

distance between two successive peaks

A

wavelength (λ)

32
Q

the number of peaks passing through a point in a second

A

frequency (v)

33
Q

range of light wavelengths

A

Shorter wavelength, higher frequency

UV 10-380
Visible 380-780
IR 780 - 300,000
Microwave 300,000 - 1,000,000,000

longer wavelength, lower frequency

34
Q

change in spectral band position in the absorption spectrum of a molecule to a longer wavelength (lower frequency); red shift

A

Bathachromic shift

*think batha = bad = red
long = bad = red

35
Q

change in spectral band position in the absorption spectrum of a molecule to a shorter wavelength (higher frequency); blue shift

A

Hypsochromic shift

36
Q

for a defined path length (b) the transmitted intensity (I) decreases exponentially with the increase in concentration of the solution (c)

A

Beer’s Law

37
Q

at a given concentration (c) the transmitted intensity of light (I) decreases exponentially with increased path length (b)

A

Lambert’s Law

*think transmission decreases with length = L = lambert

38
Q

kcb= log⁡ of Io/I

k = molar absorptivity
c = concentration
b = path length
l = transmitted intensity

A

Beer-Lambert Law

(measuring absorbance)

39
Q

In spectrophotometry, a device based on separating capability of refraction (prism) or diffraction (diffraction grating)

A

Monochromater

40
Q

Type of detector used in spectrophotometry

consists of photo emissive cathode, several dynodes, and an anode

involves multiplication of electrons that are ejected from photons that are then measured as an amplified current

A

Photomultiplier tube

41
Q

Type of detector used in spectrophotometry

can obtain info over wide range of wavelengths at one time

A

Photodiode Array (DAD):

42
Q

A low molecular weight substance that can induce an immune response when coupled to high molecular weight immunogenic molecules.

A

Hapten

43
Q

True/False:

A hapten will not stimulate an immune response

A

True

(must be coupled)

44
Q

The portion of the antibody that contains the peptide sequences that form the antigen binding site.

A

the FAB region

45
Q

Hapten + Immunogenic Molecule =

A

Immunogen

46
Q

A substance that stimulates an animal lymphocyte to produce an antibody that specifically binds to it

A

Antigen

47
Q

True or False:

The Hook Effect can produce a false positive or an inaccurately high result.

A

True

48
Q

True or False:

In an EMIT immunoassay, more drug = increased NADH production.

A

True

*Think homogenous so result is proportional to amount of drug in sample

49
Q

This can produce false positive FPIA results.
Blood proteins
Iron bound to hemoglobin
Bile salts
Lipids
Amines

A

Bile Salts

50
Q

True or False:

High concentrations of diphenhydramine cause false-positive results in some urine PCP assays.

A

True

51
Q

90% of Ig in the serum is:
IgM
IgG
IgD
IgE
IgA

A

IgG

In normal serum, about 80% is IgG, 15% is IgA, 5% is IgM, 0.2% is IgD and a trace is IgE.

52
Q

The methamphetamine antibody binds methamphetamine in a sample more strongly than the labeled amphetamine. This is an example of:
Competitive binding
Non-cooperative binding
Cooperative binding
Non-competitive binding

A

Cooperative Binding

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