Direct Detection Methods Flashcards
(122 cards)
1
Q
ENUMERATE THE ROLE OF MICROSCOPY
A
To determine presence of an infection
To characterize organisms grown in culture
To visualize etiologic agents
To screen for specimen adequacy
2
Q
is used to check if sputum is adequate for culture, whether it is just pure mucus or actually sputum itself
A
Bartlett’s classification
3
Q
materials known or reasonably expected to contain a pathogen
A
Etiologic agents (infectious substances),
4
Q
These are examples of?
(i.e., bacteria, yeast, parasites) not readily seen by the naked eye alone
A
Etiologic agents (infectious substances),
5
Q
A visible light is passed through the specimen – specimen’s image appears dark against a brighter background
A
BRIGHT-FIELD (LIGHT) MICROSCOPY
6
Q
extent to which the focused object is maintained in microscopy
A
Resolution
7
Q
the closest distance between two objects that allows to be distinguished from each other when magnified in microscopy
A
Resolving Power
8
Q
Allows the object to stand out from the background. Commonly achieved thru staining techniques
A
Contrast
9
Q
is what is used to visualize bacteria. It is also the only objective lens that requires a substance to visualize slides, as the substance (oil) prevents the scattering of light to visualize the outlines of the specimen
A
The Oil Immersion Objective
10
Q
Has no specific distance as it is dependent on your own vision
A
Interpupillary Distance
11
Q
The ability of the microscope to not lose focus when switching objectives
A
Parfocal
12
Q
Direct observation of unstained specimens – wet mount / wet preparation
A
PHASE-CONTRAST MICROSCOPY
13
Q
Direct examines of organism , NO STAINING
A
Wet Mount
14
Q
Uses a SPECIAL CONDENSER is used with an annular (ring-shaped) diaphragm – direct and reflected/diffracted light rays are brought together to produce the image
A
PHASE-CONTRAST MICROSCOPY
15
Q
Light from below the specimen is blocked (condenser), so that it reaches the object at a sharp angle – specimen appears luminous against a dark background
A
DARKFIELD MICROSCOPY
16
Q
Used primarily to view spirochetes
A
DARKFIELD MICROSCOPY
17
Q
Microscopy limited to specialized research facility
A
DARKFIELD MICROSCOPY
18
Q
Absorb light in UV range and emit visible light at a longer wavelength (when returning to their normal/lower energy state
A
Fluorochromes / Fluorophores (fluorescent dyes)
19
Q
Direct chemical interaction with the component of the bacterial cell (i.e., Acridine orange, Auramine, Calcofluor white)
A
Fluorochroming
20
Q
antibodies are conjugated (monoclonal antibodies) to a fluorescent dye (i.e., Fluorescein isothiocyanate / FITC
A
Immunofluorescence
21
Q
Fluorescing object appears bright against a dark background
A
FLUOROSCENT MICROSCOPY
22
Q
PART OF FLUOROSCENT MICROSCOPY
prevents the excitation wavelengths from damaging the observer’s eyes; selects and limits the wavelength of transmitted light
A
Excitation / Barrier filter
23
Q
often used to maintain permanent records in FLUOROSCENT MICROSCOPY
A
Digital photography
24
Q
IN FLUOROSCENT MICROSCOPY
It is the excitation light that activates the Fluorochromes, which result in the production of energy
TRUE OR FALSE
A
TRUE
25
The process of coloring the microorganism prepared in a thin film on a slide known as smear (sample)
STAINING
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smear prepared directly from patient sample / specimen
DIRECT SMEAR
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smear prepared from grown cultures in artificial media (preferably from 24-hour cultures)
Indirect smear
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Identify the smear being described
Checks slow-growing organisms
Used during Antibiotic treatment
Done when specimen is not processed immediately or
appropriately
Indirect smear
29
2 types of differential staining
Gram Staining and Acid Fast Staining
30
Principal stain utilized in bacteriologic examination
Gram Staining
31
Why is Gram Staining performed
To identify the causative agent may it be gram +, gram -, or even yeast.
32
Provides rapid presumptive identification of pathogens and assessment of specimen quality
Gram Staining
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In Gram Staining they retain the stain based on the characteristics of their cell wall
Bacteria
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Cell walls that retain the crystal violet-iodine (CV-I) complex
Thick cell walls (peptidoglycan
35
is commonly negatively charged so all stains used to visualize them are most likely basic dyes at Ph 7
Bacteria
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Primary stain: stains all bacteria blue to purple
Crystal Violet
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Mordant: enhances reaction between cell wall and primary stain
Gram's Iodine
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Decolorizer: gram-positive bacteria retain the primary stain because of the peptidoglycan and teichoic acid cross-links. Gram-negative bacteria lose the primary stain because of the large amount of lipopolysaccharide in the cell wall
Ethyl alcohol or acetone
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Counterstain: no effect on gram-positive bacteria; stains gram-negative bacteria pink to red
Safranin O or carbolfuchsin
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is the most critical step in gram staining
DECOLORIZATION
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May result in false gram –
Overdecolorization
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May result in false gram +
Underdecolorization
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Mordants like gram iodine enhances the binding of crystal violet to thick cell walls of bacteria
TRUE OR FALSE
TRUE
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Debris like proteins, squamous epithelial cells, polymorphonuclear cells, WBC’s, etc… will stain Gram – (red). Usually Gram + is reserved for yeast and other Gram + organisms
TRUE OR FALSE
TRUE
45
Quantitate the organism in Gram Stain Reporting:
10-20 per oil immersion field
Many (4+)
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Quantitate the organism in Gram Stain Reporting:
6-10 per oil immersion field
Moderate (3+)
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Quantitate the organism in Gram Stain Reporting:
3-5 per oil immersion field
Few (2+)
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Quantitate the organism in Gram Stain Reporting:
Fewer than 10 identified on complete smear
Rare (1+)
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Quantitate the organism in Gram Stain Reporting:
None were seen
None
50
Quantitate the cells (WBC’s, RBC’s and epithelial cells) in Gram Stain Reporting:
25 or more per low-power field
Many (4+)
51
Quantitate the cells (WBC’s, RBC’s and epithelial cells) in Gram Stain Reporting:
10-25 per low-power field
Moderate (3+)
52
Quantitate the cells (WBC’s, RBC’s and epithelial cells) in Gram Stain Reporting:
2-10 per low-power field
Few (2+)
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Quantitate the cells (WBC’s, RBC’s and epithelial cells) in Gram Stain Reporting:
Fewer than 2 per low-power field
Rare (1+)
54
Quantitate the cells (WBC’s, RBC’s and epithelial cells) in Gram Stain Reporting:
None were seen
None
55
Designed for bacteria (i.e., mycobacteria) whose cell walls contain long-chain fatty acids (i.e., mycolic acid) – resistant to decolorization with acid-alcohol due to the presence of Mycolic acid
ACID FAST STAINING
56
Acid Fast Staining that utilizes heat to allow penetration of primary stain into the cell wall
Ziehl-Neelsen (Classic/Hot method)
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Acid Fast Staining that replaced the heating step with a higher concentration of phenol in the primary stain
Kinyoun (Cold method)
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Acid Fast Staining that replaced the conventional acid alcohol concentration to 1% H2SO4 (intestinal coccidian oocysts). The common concentration for decolorizers is 3%
Modified Kinyoun
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These are only partially acid-fast species
Nocardia spp. And Cryptosporidium
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It just takes a speck of red to tell if a certain bacteria is Acid-fast-positive
TRUE OR FALSE
TRUE
61
In Acid fast staining we use distilled water to avoid the contamination of tap water bacillus in the specimen
TRUE OR FALSE
TRUE
62
Methylene blue, Carbolfuchsin, Crystal violet, Safranin
These are all simple or differential stains?
SIMPLE STAINS
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Utilized for observation of presence of microorganism and their size, shape, and morphology
SIMPLE STAINING
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Contains metachromatic granules stained by (methylene blue)
Corynebacterium diphtheriae
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Stain used in Negative Staining
India Ink / Nigrosin Dye
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Staining the background with an acidic dye that make cells become colorless as they repel the acidic stain (light colored bodies against a dark background)
What is this type of stain?
NEGATIVE STAINING
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What staining is being described here?
o Demonstration of diffuse capsule
o Can be utilized in the study of bacterial gas vacuoles and
viral morphology
NEGATIVE STAINING
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These are stains used in what staining category?
Anthony’s, Hiss’s and Gin’s (Crystal violet)
Nigrosin (Nigrosin dye)
Welch (Crystal violet)
CAPSULE STAIN
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Under capsule staining what undergoes this process?
Treatment with hot crystal violet solution and followed by a copper sulfate solution rinse
Welch (Crystal violet)
70
These are stains used in what staining category?
Gray (Carbol fuchsin & Tannic acid)
Leifson (Carbol fuchsin, Tannic Acid & Methylene blue)
FLAGELLA STAIN
71
Under flagella staining what undergoes this process?
Unstable colloidal suspension of tannic acid salts are used to form precipitate on the cell walls and flagella, increasing its diameter and allows staining
Leifson (Carbol fuchsin, Tannic Acid & Methylene blue)
72
These are stains used in what staining category?
Fuelgen (Carbol fuchsin)
Acridine orange
NUCLEIC ACID STAIN
73
Under nucleic acid staining what undergoes this process?
o Binds all nucleic acid
o Fluorochrome
o Useful in staining mycoplasma
Acridine orange
74
Under nucleic acid staining what is specific for DNA?
Fuelgen (Carbol fuchsin)
75
These are stains used in what staining category?
Dorner (Carbol fuchsin & Nigrosin dye)
Schaeffer-fulton (Malachite green & Safranin)
ENDOSPORE (SPORE) STAIN
76
Under endospore (spore) staining what undergoes this process?
o Observation of intracellular refractile bodies
o Heating and alcohol treatment allows penetration and
decolorization of stain
Schaeffer-fulton (Malachite green & Safranin)
77
These are stains used in what staining category?
Auramine-rhodamine
Calcofluor white
FLUORESCENT STAIN
78
Under fluorescent staining what undergoes this process?
o Binds to the mycolic acid found in mycobacteria
Auramine-rhodamine
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Under fluorescent staining what undergoes this process?
o Used in visualizing yeast
o Binds to the cell wall of fungi
Or their chitin / cellulose
o May also be used to visualize microsporidia
Calcofluor white
80
These are stains used in what staining category?
Lactophenol cotton blue
Methenamine silver
Periodic acid-Schiff
FUNGAL STAIN
81
Under fungal staining what undergoes this process?
Made up of lactic acid (preservative), phenol (inhibitory to other fungus), cotton blue (stain–chitin)
Lactophenol cotton blue
82
Under fungal staining what undergoes this process?
Stain fungal elements found in tissue samples
Methenamine silver
83
Under fungal staining what undergoes this process?
o Preferred stain for direct clinical material
o Stain molecules high in carbohydrate content
Periodic acid-Schiff
84
The study of the components and functions of the immune system
o Immune cells, WBC’s, antibodies, compliments
IMMUNOLOGY
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The study of disease diagnosis using antibody measurements in patient’s serum
SEROLOGY
86
What is being described?
proteins/polysaccharides
recognized by the immune system
found in cell walls as epitopes
Antigen
87
molecule capable of eliciting an immune response
Immunogen
88
products of the immune response following interaction with an immunogen
Antibody
89
All immunogens are antigens, because all antigens can elicit an immune response
TRUE OR FALSE
FALSE
All immunogens are antigens, but not all antigens can elicit an immune response
90
Aggregation of soluble antigens and antibodies
Precipitation
91
These are under what concept of precipitation?
Radial immunodiffusion
Subjective interpretation and low sensitivity
Single Immunodiffusion / Oudin gel diffusion
92
Single Immunodiffusion / Oudin gel diffusion if done in a petri dish
Radial immunodiffusion
93
Commonly used in the detection of exoantigens
Double Immunodiffusion / Ouchterlony gel diffusion
94
These are examples of?
Systemic fungi (i.e., Blastomyces dermatitidis, Coccidioides immitis, Coccidioides posadasii, Aspergillus spp., Histoplasma capsulatum) and Candida
Exoantigens
95
What is being described?
Most widely-used in the detection of T. pallidum
Venereal Disease Research Laboratory / VDRL
Rapid Plasma Reagin / RPR
Flocculation
96
Clumping or aggregation of cells or coated particles toward a specific antigen / antibody
Agglutination
97
Agglutination wherein Natural carrier molecules (i.e., directly on organism’s surface)
Direct Agglutination
98
Identify this type of Agglutination
Identifies antigens rather than antibodies
Antigen bound to a variety of particles (i.e., latex beads,
gelatin, erythrocytes)
Indirect / Passive agglutination
99
Commonly used in detection of:
Cryptococcus neoformans in CSF / Serum, Clostridium
difficile toxins A and B
Presence of beta-hemolytic streptococci, antibodies to
viruses (i.e., rotavirus, CMV, rubella, VZV)
Indirect / Passive agglutination
100
Febrile agglutinin test, Widal Test, Weil-Felix Test are all test under what agglutination?
Indirect / Passive agglutination
101
What type of agglutination is being described?
Antibody bound to a carrier particle instead of antigen
Commonly used in rapid identification of antigens from
Group A & B streptococcus, Staphylococcus aureus,
Cryptococcus neoformans
Reverse Passive agglutination
102
Agglutination of Carrier molecules
Coagglutination
103
Agglutination that uses a killed or treated Staphylococcus aureus organisms (Cowan I Strain) and Antigen-antibody binding protein (Protein A)
Coagglutination
104
Agglutination used commonly used in Identification of Lancefield Groups (A, B, C, D, F, G, N), Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae (A-F)
Coagglutination
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Utilizes antibody (monoclonal / polyclonal) conjugated to a fluorochrome (FITC)
IMMUNOFLUORESCENCE
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Antibody conjugated to a fluorochrome added directly with the patient antigen fixed on the slide
Direct Fluorescent Antibody
107
Immunofluorescence commonly used in detection of antigens toward Legionella pneumophila and Chlamydia trachomatis
Direct Fluorescent Antibody
108
More commonly used than Direct Fluorescent Antibody (DFA)
Indirect Fluorescent Antibody
109
Patient antibody is incubated with a known antigen attached to a solid phase and is then washed and added with anti-human immunoglobulin conjugated to a fluorochrome
Indirect Fluorescent Antibody
110
IMMUNOFLUORESCENCE Commonly used in Legionella spp., Borrelia burgdorferi, T. gondii, VZV, CMV, EBV capsid antigen & nuclear antigen, HSV 1 & 2, rubella, Mycoplasma pneumoniae, T. pallidum, and several rickettsiae
Indirect Fluorescent Antibody
111
is commonly used to identify Treponema pallidum seen in patients screening for syphilis
Indirect Fluorescent Antibody
112
May also be called Enzyme Immunoassay (EIA) and Described into two competitive and non-competitive
ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA)
113
Antigen / antibody bound to an enzyme – utilization of enzymes as labels
ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA)
114
ELISA where the Antibody is attached to a microdilution tray, spherical plastic / metal bead
Solid-phase
115
ELISA commonly used in detection of HBsAg & early hepatitis B antigens, HIV p24 protein
Solid-phase
116
ELISA that is Flow-through and large surface area (absorbent material pulls the reactants through the membrane) –nitrocellulose, nylon
Membrane-bound solid phase
117
ELISA commonly used in detection of antibody towards hepatitis virus, HSV 1 & 2, RSV, CMV, HIV, rubella (IgG & IgM), mycoplasmas, chlamydiae, B. burgdorferi, Entamoeba histolytica
Membrane-bound solid phase
118
These are what kind of ELISA?
Slot-blot, Dot-blot assays, Cassette-based membrane-bound ELISA
Innovations
119
An assay used for detection of inhibitory effects of antibody to viruses, BUT is replaced by particle agglutination tests
NEUTRALIZATION ASSAY
120
Assay commonly used in detection of antibodies toward bacterial toxins, extracellular products (i.e., Streptolysin O, Dnase B)
Generally performed at research laboratories
NEUTRALIZATION ASSAY
121
Assay that uses demonstration of antibody presence at the patient sample
COMPLEMENT FIXATION ASSAY
122
Assay commonly used in detection of antifungal and antiviral antibodies, BUT is replaced by: IFA and ELISA
COMPLEMENT FIXATION ASSAY
