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Laboratory Diagnostics Flashcards

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1
Q

What can erythrocytes by indications of?

A

Anemia and erythrocytosis

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2
Q

What can leukocyte levels be indicators of?

A

Inflammatory conditionsNeoplastic conditionsChemotherapy

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3
Q

What can platelet levels be indicators of?

A

Bleeding disordersDisseminated intravascular coagulation

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4
Q

How are counts and morphology carried out in the laboratory?

A

Counts done on different cell types by machineMorphology checked by microscope

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5
Q

What is the packed cell volume?

A

Distance of buffy coat divided by total distance

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6
Q

What is the buffy coat?

A

White blood cells

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7
Q

What colour should plasma be and what would cause a colour change?

A

Clear or straw coloured normallyPink if hemolysed

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8
Q

What would contraction of the spleen cause to be dumped into the blood stream?

A

A large amount of red blood cells

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9
Q

What is contained in a complete blood count?

A

RBC concentration - no. of red blood cellsHGB - total haemaglobinHCT - haematocrit and spun PCVMCV - mean cell volumeMCH - mean cell haemoglobinMCHC - mean cell haemoglobin concentrationRDW - red cell distribution width

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10
Q

What should total haemoglobin usually correspond to?

A

Red blood cell concentration

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11
Q

What does the mean cell volume tell you?

A

How big red blood cells are

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12
Q

What is the mean cell haemoglobin concentration?

A

How much haemoglobin there is per red blood cell (mg/volume of RBC)Red cells mature when they reach a certain concentration of haemoglobin

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13
Q

What are the 3 classifications of anemia?

A

Mil, moderate or severe - can give an idea of underlying problem

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14
Q

What features can be used to determine regenerative anemia from non-regenerative?

A

MCV - increased during regenerative as reticulocytes are larger than mature erythrocytes - normal in non-regenerativeMCHC - decreased in regenerative anaemia as reticulocytes are larger and less packed with haemoglobin - normal in non-regenerative

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15
Q

When are microcytic red blood cells seen and why?

A

During portosystemic shunt, iron deficiency and hepatic failureIron is needed to build haemoglobin so with less iron get less haemoglobin so RBC divide again as a certain Hb concentration is needed to become more mature

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16
Q

Which breed of dog is microcytic anaemia a normal thing to see?

A

Akitas - born with smaller RBC

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17
Q

When are macrocytic red blood cells seen?

A

In regeneration - polychromatophils are larger than mature cells

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18
Q

Which breed of dogs can have a rare case where all RBC are larger?

A

Poodles

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19
Q

What do hypochromic cells look like?

A

Lots of cells without Hb concentration

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20
Q

Why are hyperchromic cells not seen unless artificially?

A

Haemolysis has to occur for them to be seen - not possible naturally

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21
Q

What is the first thing we should classify when identifying anaemia?

A

Whether it is regenerative or non-regenerative

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22
Q

What are the only 2 reasons for regenerative anaemia?

A

Haemolysis (destroying blood)Haemorrhage (losing blood)

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23
Q

What generally appears with regenerative anaemia?

A

Larger RBC appearing as bone marrow isn’t damaged and they still pump out RBCs

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24
Q

What are the three most common non-regenerative causes of anaemia?

A

Anaemia of inflammatory/chronic disease - mildChronic renal failure - no EPO being producedDecreased production in marrow - not producing RBC

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25
Q

What is the difference between reticulocytes and polychromatophils?

A

Same cell but different - only reticulocytes when RNA is precipitated

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26
Q

How can we see polychromatophils?

A

Quik or Giemsa stained smear - younger cells containing ribosomal RNA show up as larger, bluer cells

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27
Q

What do we use to see reticulocytes on a blood smear?

A

Stain polychromatophils with New Methylene Blue and RNA precipitates forming aggregates - reticulocytes

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28
Q

What, in cats, is the same as reticulocytes in other animals?

A

Aggregate retics - do mature to punctate retics over time but only really interested in aggregate retics

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29
Q

What does an increase in reticulocyte percentage mean and why can it be misleading?

A

More young red blood cells meaning anaemia is regenerativeCan be misleading if red blood cell count is not normal

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30
Q

What is the corrected reticulocyte percentage?

A

(Reticulocyte percentage multiplied by patient PCV)/normal PCV

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31
Q

What are the normal corrected reticulocyte percentages for a dog or a cat?

A

45% for dogs35% for cats

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32
Q

When can anaemia be categorized as regenerative when regarding corrected reticulocyte percentages?

A

> 1% corrected in dogs>0.4% corrected in cats

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33
Q

What signs of regeneration can be seen on a blood smear?

A

Polychromasia - high number of red blood cellsAnisocytosis - variability of RBCMacrocytosis - enlarged RBCnRBCs - nucleated RBCHowell-Jolly bodies - remains of nucleus (more in regen.)Codocytosis - cells with bullseye appearanceBasophilic stippling - small periphery dots

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34
Q

What does a blood smear look like with a lack of regeneration?

A

Uniform cells - pretty uneventful

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35
Q

What 5 things should be looked for when looking at RBC morphology?

A

Spherocytes/ghost cellsHypochromasia/leptocytosisShear productsOxidative damageOrganisms

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36
Q

What should be looked for in a total white blood cell count?

A

NeutrophilsLymphocytesMonocytesEosinophilsBasophils

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37
Q

What should the typical differential of leukocytes be?

A

80-95%

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38
Q

Where do neutrophils spend most of their time and how do they travel?

A

In tissues - travel in bloodstream

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39
Q

If different counts are taken of neutrophils throughout the day what will the values look like?

A

Values will differ

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40
Q

What is an increase in neutrophils called?

A

Neutrophilia

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41
Q

What is a decrease in the total number of neutrophils?

A

Neutropenia

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42
Q

What is left shift of neutrophils?

A

A bigger demand for neutrophils so younger cells begin to come into the circulation

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43
Q

What is the difference between regenerative left shift and degenerative left shift?

A

Regenerative - neutrophilia - segmented>bandsDegenerative - neutropenia - bands>segmented

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44
Q

What is leukemia?

A

Neoplastic cells within the circulation

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45
Q

Which lype of leukemia is worse?

A

Acute - will kill quicklyChronic - bad in long term but in short term they’re alright

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46
Q

Describe an acute leukemia

A

Blast cells in circulation - can’t identify cell of originMore likely to be lymphoidIf segmentation - myeloidDetect by immunophenotyping using a flow cytometer

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47
Q

Describe chronic leukemia

A

Lymphoid - persistent high numbers of small, mature lymphocytesMyeloid - persistent high numbers of normal neutrophils

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48
Q

What is a common problem with platelet measurements?

A

Clumping due to activation of ability to plug holesMachine can’t distinguish platelet from clump

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49
Q

What are five examples of clinical pathology tests?

A

HaematologyClotting profileBiochemistryUrinalysisCytology

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50
Q

What is biochemistry used for?

A

Evaluating different organ systems - liver, kidney etc.

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51
Q

What should biochemistry be used with?

A

Urinalysis and CBC

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52
Q

What is recommended most of the time for biochemistry?

A

Serum not plasma

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53
Q

What should be used for biochemistry for birds and reptiles and why?

A

Heparinized plasma - small sample size

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54
Q

What is the difference between serum and plasma?

A

Plasma contains all clotting factorsSerum is fluid part of blood after clot formation

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55
Q

Describe serum collection

A

Collect into tubes without anticoagulantUsually have a red or brown topAlso have serum separator tubes with gel layer separating cells and serum

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56
Q

Why do we seperate red cells and serum?

A

Red cells continue living and can undergo changes so seperate so no more interface between serum and RBC

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57
Q

How is serum seperated?

A

Allow blood to clot and centrifuge - should be done immediately if possible

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58
Q

What temperature should serum be stored at?

A

4 degrees C

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59
Q

How is plasma collected?

A

Blood collected into EDTA, heparin or citrateSeparate using centrifugeStore at 4 degrees C

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60
Q

Give at least 5 examples of things found in a biochemistry profile

A

Total proteinAlbuminGlobulinsBicarbonateAnion gapCalciumPhosphorusGlucoseElectrolytesUrea nitrogenCreatinineBilirubinCholesterolAmylaseLipaseCK - creatine kinaseALT - alanine transaminaseALP - alkaline phosphataseSDH - saccharopine dehydrogenaseGLDH - glutamate dehydrogenase

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61
Q

Why should caution be taken when looking at an animals biochemistry profile?

A

Only 95% of animals fall within reference rangesLarge number of things measuredLikely that some of their values fall outside these rangesCould be a problem but sometimes mean nothing

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62
Q

What are alterations to levels of chemicals in the blood always a result of?

A

Either a change in the amount going in or the amount going out

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63
Q

Describe how total protein is measured and what it is made up of?

A

Measured on serum or plasma (slightly greater due to fibrinogen) by refractometer or colorimetric methodMade up of albumin and globulins

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64
Q

What are total protein levels falsely increased by?

A

Icterus, severe haemolysis and lipemia

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65
Q

What can caused increased protein levels?

A

DehydrationInflammationNeoplasia

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66
Q

What can cause decreased protein levels?

A

Blood lossDecreased synthesisDilution

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67
Q

How is albumin measured and in what animals is this method unreliable? What method should be used instead?

A

Dye binding methodUnreliable in birds - use electrophoresis instead

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68
Q

When is increased albumin seen? Decreased?

A

Increased - dehydrationDecreased - increased blood loss, decreased synthesis and third spacing

69
Q

How can the different types of globulin be separated?

A

Electrophoresis

70
Q

What can cause a selective increase in globulins?

A

Inflammation (polyclonal)Neoplasia (monoclonal)

71
Q

What is the difference between polyclonal and monoclonal gammopathy?

A

Polyclonal - increase in all globulin typesMonoclonal - increase in a single globulin type by a single clone of cells

72
Q

What three things can be used to evaluate the liver?

A

EnzymesMetabolitesFunction tests

73
Q

What can be used to evaluate pancreas function?

A

Amylase and lipase

74
Q

Why are measurements of amylase and lipase not specific indicators of pancreatic problems?

A

Can have increases without pancreatic damage e.g. kidney damage

75
Q

When are increases in amylase and lipase seen in dogs and cats?

A

Dogs = 4 to 5 fold increase with pancreatitisCats = not seen with pancreatitis

76
Q

What can cause increases in amylase and lipase?

A

Renal insufficiencyPancreatitis - dogs

77
Q

What do we need to look at to evaluate the urinary system?

A

Both serum/plasma and urine

78
Q

What are both indicators of glomerular filtration and where are they produced?

A

Urea - produced in liver and excreted by kidneyCreatinine - derived from creatinine in muscle

79
Q

What else can the urinary system be used as a marker for?

A

Liver and kidney function

80
Q

What is azotemia? What can cause it?

A

Increases in nitrogenous wastes in the circulationCaused by dehydration, renal disease and obstruction

81
Q

What should be checked to determine azotemia?

A

Urine specific gravity - ability to concentrate urine so should compare with serum/plasma urea and creatinine

82
Q

What should urine specific gravity be if urea and creatinine are increased?

A

1.030 in dogs1.035 in cats1.025 in horses/ruminants

83
Q

What is happening if urine specific gravity is less than the recommended values?

A

Decreased concentrating abilityRenal failure is present

84
Q

What isn’t a reliable indicator of renal disease?

A

Urea - should use creatinine only

85
Q

What are urea levels influenced by and why may they be increased?

A

Influenced by protein intakeIncreased by high protein meal or GI bleeding

86
Q

Why is urea not a reliable indicator of renal disease in ruminants?

A

Secrete into the saliva and goes into the rumen

87
Q

What is the main ion in extracellular fluid?

A

Sodium

88
Q

What regulates sodium along with water?

A

Kidney

89
Q

How can increased sodium occur? Decreased?

A

Increased - increased intake or water loss, decreased water intakeDecreased - increased loss or water intake

90
Q

What is the main ion in the intracellular space? What affects its levels?

A

Potassium - affected by acid-base changes

91
Q

What regulates potassium levels? What causes increases or decreases?

A

Regulated by intake and renal functionIncreases - renal failure and hypoadrenocorticism and cell leakageDecreases - loss and decreased intake

92
Q

What should be considered before investigating an in/out problem with potassium?

A

Whether an acid-base balance shift that would cause potassium shift

93
Q

What do changes in chloride usually coincide with?

A

Sodium

94
Q

What are changes in chloride without sodium changes associated with?

A

Alteration in acid/base statusVomiting/abomasal displacement

95
Q

What two states is calcium present as within the body?

A

Free - activeBound - bound to albumin

96
Q

What is calcium affected by?

A

Albumin levels

97
Q

What are calcium levels regulated by?

A

PTH and calcitonin

98
Q

What is phosphorous regulated by?

A

PTH and calcitonin

99
Q

What are increases in phosphorous associated with and when are they often seen?

A

Increase associated with renal diseaseSeen in growing animals along with elevated calcium and ALP

100
Q

What three things are looked at in urinalysis?

A

Gross appearanceChemical analysis - specific gravity and urine stripSediment analysis - cellular elements, crystals, casts and others

101
Q

What are the advantages of cytology?

A

Quick, easy and inexpensiveNon-invasiveMinimal risk to patientCan determine what diagnostic procedure should be performed next

102
Q

What are some limitations of cytology?

A

Relies on sample quality - skill of collector, smear quality and tissue typeLimited by ability/experience of person examiningLack of information about tissue architectureDiagnostic challenges

103
Q

What are the advantages and disadvantages of histopath?

A

-More expensive- Slower turn around time- Poor detail for round cell tumours+Tissue architecture+Tumour grading+Immunohistochemistry more available

104
Q

What are the two types of samples used for cytology?

A

Aspiration or imprints- superficial masses, lymph node and organs/deep massesFluids- body cavities, joints, respiratory tract and cerebrospinal fluid

105
Q

What should fine needle biopsies be used for?

A

Solid or fluid filled masses

106
Q

What should fine needle biopsies be done under?

A

Visual or ultrasound guidance

107
Q

Describe what is done during a fine needle biopsy

A

Similar to fine needle aspiration but no negative pressure is applied to the syringeSmall gauge needle (22-24 gauge) inserted into mass several times - also sample wall not just centreUse needle to take sample then air-filled syringe to expel cells onto slide

108
Q

What is the difference between a fine needle aspiration and a fine needle biopsy?

A

Aspiration uses negative pressure to draw sample from a mass redirecting needle 2-3 times to ensure a representative sampleBiopsy doesn’t use negative pressure i.e. drawing a syringe back

109
Q

What needs to be done to ensure cells can be seen on a smear?

A

Flatten the cells out

110
Q

Describe smear preparation

A

Remove needle from syringeDraw air into the syringeReplace needleExpel aspirated fluid onto slide being gentleSmear - don’t spray

111
Q

What are the three goals with smear preparation?

A

Thin areas with good cell spreadMinimization of cell damageMinimization of blood content

112
Q

Describe how to make smears?

A

Put another slide over the top of the samplePull slides apart - use gentle pressure

113
Q

What happens if too much or too litle pressure is used when making a smear?

A

Excessive pressure - rupture cellsNot enough pressure - too thick preparation

114
Q

What are touch impressions good for?

A

Evaluation of excised tissue or superficial lesions

115
Q

When are imprints made?

A

Before excised tissue is placed in formalin and submitted for histopathology

116
Q

Describe how to take an imprint

A

Use fresh cut surface of the tissueBlot until drywith paper towelImprint directly onto glass slide - roll 4 to 5 times then air dry and stain

117
Q

What normal cells do we expect in various samples?

A

Tracheal wash - ciliated cellsBAL - alveolar macrophagesJoint fluid - mononuclear cells

118
Q

What is the difference between inflammation and neoplasia in cytology?

A

Inflammation - sample dominated by inflammatory cellsNeoplasia - sample dominated by tissue cells

119
Q

Describe the differences between septic and non-septic inflammation upon cytology

A

Septic- contains bacteria/organisms, degenerate neutrophils, bacteria must be intracellular within neutrophils to be significant also may be contaminantsNon-septic- no bacteria or organisms seen and non-degenerate neutrophils present

120
Q

Describe degenerative changes in neutrophils

A

Nuclear changeNucleus swells, loses lobulation and becomes palerSecondary to release of bacterial toxins

121
Q

When are increased numbers of macrophages seen in cytology?

A

Granulomatous inflammationPyogranulomatous inflammation - if neutrophils alsoForeign body reactions

122
Q

Which tumours present as round cell tumours?

A

LymphomaHistiocytomaMast cell tumourPlasmacytomaTransmissible venereal tumourMelanoma

123
Q

Describe epithelial cells upon cytology

A

Sheets/rafts/clustersLarge cell sizeCell-to-cell junctionsOval to angular in shapeNuclei round, centrally locatedCytoplasm often abundantGood cell yield

124
Q

Describe mesenchymal cells upon cytology

A

Individual cells or clumpsSmall to medium size cellsSpindle to fusiform to stellateIndistinct cell bordersElongated nucleusPoor exfoliationProduced in the matrix

125
Q

How can malignancy be assessed?

A

Uniformity vs. pleomorphismMonotonyNuclear criteria are the most reliable

126
Q

What is needed before you can call a tumour malignant?

A

Need at least three nuclear criteria of malignancy

127
Q

What are the three cellular criteria of malignancy?

A

Anisocytosis - variation in cell sizeMacrocytosis - large cellsCell crowding

128
Q

What are nuclear criteria of malignancy?

A

Anisokaryosis - variation in nuclear sizeMultinucleation - nuclei vary in size and odd numbersMacrokaryosis - giant nucleiHigh nuclear to cytoplasmic ratioIncreased mitotic figuresAbnormal mitotic figuresCoarse chromatinNuclear moulding - nuclear deformation by other nucleiMacronucleiVarying nucleolar shapeVarying nucleolar size

129
Q

What are some common problems with cytology samples?

A

Formalin fumes - destroy blood or cytologyRefrigerating glass slidesBreakage during shippingLack of freshly made smears

130
Q

What are the two immunodiagnostics used in veterinary medicine?

A

Serology and immunoassay

131
Q

What does serology allow?

A

Evaluation of immune status/function - exposure of animal to infectino, response to vaccination and diagnosis of immune-mediated disease

132
Q

What does immunoassay allow in veterinary medicine?

A

Use of labelled antibodies as detection reagents - presence of pathogen in a sample, measurement of a biomarker and immunophenotyping

133
Q

What two ways can we sample the immune system?

A

Blood sample and tissue biopsy

134
Q

What are the two different types of blood sample you can take?

A

Serum - clotted sampleCells - citrate/heparin anticoagulant

135
Q

What are the serological markers of innate immunity?

A

Acute phase proteins - C-reactive protein, serum amyloid A, fibrinogen and haptoglobin

136
Q

What are the serological markers of adaptive immunity?

A

Antibodies and cytokines

137
Q

What two ways can be used to measure total immunoglobulin?

A

Radial immunodiffusion and serum protein electrophoresis

138
Q

What three things can measurement of total immunoglobulin be useful in?

A

Failure of passive transfer in foalsSpecific Ig deficiency syndromesMonoclonal/polyclonal gammopathy

139
Q

What can antigen-specific immunoglobulin measurement be useful in determining?

A

Exposure of an antigen to pathogenResponse to vaccinationDiagnosis of antibody-mediated hypersensitivity

140
Q

Describe serum protein electrophoresis

A

Various peaks of protein levelsNormally albumin should be dominant peakCan be used to divide albumin and globulins - gamma globulin most important

141
Q

What is seen in a monoclonal gammopathy on serum protein electrophoresis?

A

One peak of protein increased

142
Q

What are the seven ways you can measure antigen-specific antibody?

A

ELISAImmunofluorescent antibody testVirus neutralisation assayComplement fixation testHaemagglutination inhibitionMicroscopic agglutination testAgar gel immunodiffusion test

143
Q

What two things can ELISA be used to detect?

A

Antigen - pathogen presenceAntibody - immune response

144
Q

What is better to look for if exposure to a pathogen was recent?

A

Presence of the causative organism in infected tissue or biological fluids

145
Q

What is it better to look for if exposure occurred greater than seven days ago?

A

Antigen specific antibody in serum

146
Q

What would you expect to see in an ELISA with a recent infection?

A

Greater IgM:IgG ratio and an increasing IgG titre

147
Q

What is different about an immunofluorescence assay compred to an ELISA?

A

Destination antibody is labelled with a fluorescent marker rather than an enzyme - fluorescence indicative of a positive result

148
Q

What is a viral neutralisationg assay more indicative of than an ELISA?

A

Presence of a biologically active antibody

149
Q

Describe the basics behind a viral neutralisation assay

A

Cultured cells deliberately infected with virus either with serum or withoutIn the absence of specific antibody cells are infected

150
Q

What is the advantage of a virus neutralising antibody over an ELISA?

A

Indicates presence of biologically active antibody

151
Q

How can T cell responses be evaluated in vitroor in vivo?

A

In vitro- cytokine release following stimulation with specific antigenIn vivo- delayed-type hypersensitivity test

152
Q

Describe the bovine TB gamma-IFN test

A

Heparinised blood sample sent to AHVLACulture cells in vitro with mycobacterial antigensMeasure IFN-gamma production by ELISAMore sensitive but less specific than tuberculin “skin test”

153
Q

What should be tested to diagnose immunodeficiency syndromes?

A

Neutrophil function

154
Q

What is measured with serology-based tests in allergic disease?

A

Allergen specific IgE

155
Q

What are the two types of hypersensitivity that can be measured with intra-dermal skin testing?

A

Immediate-type - IgE mediatedDelayed-type - T cell mediated

156
Q

What are intradermal skin tests used a lot for?

A

Diagnosis of small animal allergic skin disease and immediate-type hypersensitivity

157
Q

What are three tests used for immunodiagnostics of autoimmunity?

A

Coomb’s test for IMHA (Immune Mediated Haemolytic Anaemia)Anti-nuclear antibody (ANA) for systemic lupus erythematosus (SLE)Specific autoantibody serology

158
Q

What three things can antibodies be used as detection reagents for?

A

Presence of pathogen in biological sampleMeasurement of a biomarkerImmunophenotyping

159
Q

What three ways can antibodies be used to detect infection within a sample?

A

Sandwich ELISA (biological fluid)Immunofluorescence (biological fluid and tissue)Immunohistochemistry (tissue)

160
Q

What are the methods used for measurement of a biomarker using labelled antibodies?

A

ELISAChemiluminescenceRadioimmunoassay

161
Q

How can cell type presence be identified within a biological fluid or tissue biopsy?

A

Antibodies against cell-surface marker

162
Q

What are the two most common methods of identifying a pathogen’s nucleic acid in a biological sample?

A

PCR or qPCR

163
Q

What can molecular techniques be used for in pathogens?

A

Genotyping the pathogen or epidemiological studies

164
Q

What things need to be considered when using genetic tests of animals?

A

Insertion, deletion or substitutionInherited as simple or complex disorderAutosomal or sex-linkedDominant or recessiveCopmlete or incomplete penetrance

165
Q

What sample would you need for a molecular diagnostic assay for FIA?

A

EDTA blood

166
Q

How would you process a sample for a molecular diagnostic assay for FIA?

A

Extract DNA

167
Q

What would you need to know to develop a test for molecular diagnosis of FIA?

A

DNA sequence for the three mycoplasma spp for primers/probesCould you design a test for all or could you do tests for each?

168
Q

What type of assay would you use as a molecular diagnosis for FIA?

A

PCR or qPCRqPCR more specific and allows an estimate of pathogen load

Principles of Science (9 decks)

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