concepts of clinical testing Flashcards by jood kh

patient management uses:

Medical examination
* Clinical history
– Presentingcomplaint – Pastmedicalhistory
* Physical examination
* Differential diagnosis
* Investigations
– Laboratoryand/orRadiology
* Diagnosis
* Treatment
– SurgeryorMedical
* Monitor outcome
– Diseaseandtreatment

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labprtary investgiations uses:

Clinical Biochemistry
including endocrinology and
metabolism
Clinical Microbiology
Haematology
Immunology
Medical Genetics

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the use of biochemical technique in the study of molular basis of disease and in diagnosis and management
the aim is to provide an answer to a question posted about a patient
is ——-

clinical biochemistry

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the process of the clinical biochemistry:
1- selection of appropriate —-
2- measuring of analytes in —-
3- interpetation of —-

investigation
body fluids
results

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the 3 phases includes:
1-pre analytical: —–
2- analytical : ——
3- post analytical : —

1-Appropriate investigation & sample collection
2-
* Method selection
* Quality control & uncertainty of measurement
3- interpretation of results

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—- is the use of appropriate investigation and sample collection
-sample examples include:

pre analytical
- Blood
  Whole blood
  Plasma
  Serum - Urine
  Spot urine sample
  24-hour urine collection
Cerebrospinal fluid
Sweat
Faeces
Sputum & saliva
Tissue & cells
Aspirates e.g. joint (synovial) fluid

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1-pre analytical phase needs patient — such as:
2- the blood sample type such as:

1- patient preparation such as:
note of:
* Age of patient - Adult or child
* Time of day- Morning, afternoon or nighttime
* Fasting or post-prandial
* Exercise
* Medication
* Patient preparation
* Cleanse venepuncture site
2-:
- whole blood: serum clotted sample
- plasma : preservative , lithium heprin , EDTA
- transport of sample to lab: by porter + pneumatic tube transport
- laboratory info system (LIS): data entry
- sample preparation: labelling + centrifugation

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analytical methods needs:
and it includes

needs: method selection and faulty control & uncetentiy of measurements
in includes:
1- chromatography : GC or tandem -MS
2- immunological assays: enzyme linked immunosobenant assay ( ELISA)
3- spectrophotometric assay
4- nephelometric : light scatter

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—- is used to provide unknown compounds by determining mass and matching to known spectra
it provides:

-mass spectrometry
it provides:
* elemental composition of sample
* masses of particles and molecules
* potential chemical structures of molecules by
analyzing the fragments
* Produces spectra of masses from molecules in sample and fragments of the molecules m/z

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stages of mass spectrometry:
1- ioniser converts some of the sample into —
2- mass analyser separates ions according to their —-
3- detector records either — or —- when ions passes by or hits surface

ions
mass-to-charge ratio
charge induced or current produced

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—- is the application of science of enzymes to diagnosis of disease and monitoring of treatment and disease progression
- the measurement of enzyme activity or concentration in blood to:

clinical enzymology
1- diagnosis: diagnose disease
2- complications: determine complications and extent of disease
3- monitoring: monitor disease progression
( check slide 16)

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duchnenne musuclualr dystrophy:
- it results in a cycle of – and rapture of — in skeletal muscle
- this results in the release of the myocytes —-
- some of the myocytes cytoplasmic content including CK enzyme is released into —

growth and rapture of myocytes
cellular content
blood

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CK and the diagnosis of Duchenne muscular dystrophy:
- CK ( creatine kinase ) is an enzyme involved in —-
- presents in large amount in —- and —-found in — due routine turnover of skeletal muscle
- elevated – activity in DMD patient ( can be greater than 10x the reference range )
- elevated plasma CK activity in 75% of —- female carriers

energy production
skeletal and cardiac muscle
blood
plasma
asymptomatic

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other reasons of muscle damage includes:

Excessive physical exercise
- Marathon runners
- Recent strenuous exercise
Surgery
Skeletal muscle trauma Drug induced

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1-in cellular damage - why do we measure enzymes ?
2- disadvantages:

Relatively easy to measure compared with other proteins Can measure the enzyme product
lack of tissue specificity and many enzymes present in more than one tissue
( check slide 23)

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liver enzymes - plasma enzymes used to assess hepatic function includes :

AST and ALT - aspartate and alanine aminotransferase
(formally called transaminases and still abbreviated to AST and ALT respectively)
ALP - alkaline phosphatase
GGT - g-glutamyl transferase

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liver diseases
1- pathological changes :
2- tests for liver disease:

1- pathological changes
- liver cell damage: acute hepatitis, chronic hepatitis, toxin action, alcohol abuse
- cholestasis: gallstones, cancer at the head of the pancreas Reduced mass of functioning cells (e.g cirrhosis, terminal hepatic failure) Infiltrative disorders - liver is invaded or replaced by nonhepatic substances such neoplasm or amyloid.
2- tests for liver diseases:
-Liver cell damage - enzymes - ALT, AST
-Synthetic function - prothrombin time, albumin
-Conjugating capacity - conjugated bilirubin
-Cholestasis - enzymes - Alk phos, γ-GT (also serum bilirubin)

hepatoceullar damage:
— is more liver specific than — however both enzymes are widely distributed in the body tissue but — is present only in small amounts in liver
— has cytoplasmic and mitochondrial iosoenzymes which tends to be released more than — in chronic hepatocellular disease

— is known as the failure of normal amounts of bile to reach the intestine
causes:

cholestasis
mechanical block (gallstone, tumour); liver disease; drug induced, e.g. phenobarbitol
more info : Alkaline phosphatase (ALP) and g-glutamyl transferase (g-GT) – normally “anchored” to membranes of hepatocytes, released in only small amounts in hepatocellular damage.

in cholestasis , the synthesis of these enzymes is induced and they are made –

soluble ( high concentration of bile acids )
- g-GT more liver-specific – ALP also raised in bone disease
-Changes in g-GT and ALP often parallel each other in cholestatic disease

alcoholic liver disease
- the abnormalities depend on – of the disease
- the biochemical abnormalities include:

severity ( as hepatitis acute or chronic , cirrhosis )
AST>ALT
-Raised γ-GT
-Increased mean cell volume (MCV) -Hyperbilirubinaemia
-Raised Alkaline Phosphatase
-Increased prothrombin time (INR) -Hypertriglyceridaemia
-Decreased K+, PO43-, Mg2+

—- are the parameters indicate that liver disease is due to excessive alcohol intake

Raised γ-GT
Increased mean cell volume (MCV)

-The appearance of a small amount of enzyme in the bloodstream from damaged tissue can be detected with —-
- enzymes must be — and conditions/buffer are important

great sensitivity , active

measurement of enzymes in blood - the factors affecting enzymes activity:

Substrate concentration
Temperature of reaction
pH of buffer
Presence of cofactors
enzyme activators or inhibitors
Urea & related compounds may disrupt hydrogen bonds causing inactivation

1------ are multiple forms of an enzyme 2- tissue specific ones is when certain genes may be expressed exclusively in --- tissue as: 3- has cellular --- within specific organelles as mitochondria and cytoplasm

-izoenzymes - one -as: LD , CK , alkaline phosphatase - cellular localisation

izoenymes are multiple -- of an enzyme that posses the ability to ----- they differ in ---- example:

- multiple forms - catalyse enzymes characteristic reaction - differ structurally - encoded by district gene - example: creatine kinase and lactate dehydrogenase ( check slide 33)

--- is known as the death of part of the whole heart due to a blockage of an artery

mycocardial infraction

cardiac biomarkers as ---- are ---- as they rise and then slowly fall over a period of time after an ---

- CK-MB or troponin - temporal - infraction

measuring CK isotype levels:

* The different isotypes have different electrophoretic mobility. * These can be quantified * MM and MB forms predominantly increased after infarct. * The brain isoform is not increased significantly ( check table 38)

plasma enzyme levels dependant on :

Rate of release from damaged cells Rate of damage to cells Extent of cell damage In the absence of cell damage, rate of release depends on: - Rate of cell proliferation (malignancy) - Degree of induction of enzyme synthesis Rate of clearance (removal) from circulation

why measure enzyme activity in body fluids :

* A small increase in enzyme in the blood from damaged tissues can be detected with great sensitivity * Monitor changes in substrate or product concentrations * Easier than measuring protein directly as the signal is amplified from the reaction * Can measure co-factor changes * Enzymes are also antigens can be measured directly - bind to enzyme-specific antibodies

immunochemical technique is when -- used to detect chemical substance of interest ( antigen ) - depends of -- and high -- of antibody for specific antigen coupled w ability pf antibody to -- w antigen - allows --- and --- of specific substance by variety of methods

- antibody - sensitivity - affinity - cross link - identification - quantification

isotopic and non isotopic immunoassay:

Method of detection: -Isotopic: Scintillation -Non-isotopic: chemiluminescence, fluorescence, photometry -Safety – hazards, handling & disposal example: 1-Enzyme Immunoassays Enzyme-linked Immunosorbent assays - ELISA 2-Enzyme Multiplied Immunoassay Techniques - EMIT ( check till slide 49 pls)

enzyme multiplied immunoassay technique - EMIT: 1- homogenous assay to -- drugs , hormones , and metabolites ( antigens ) 2- patient sample added to a --- containing ---- and --- 3- antibody -- enzyme activity when no drug present 4- if there's is a drug in the patient sample , this competes for the --- 5- this --- the enzyme site allowing the -- to occur

- analzye - tube - labelled enzyme and antibody - antibody - opens - reaction

In EMIT: * No ------ required- patient sample mixed with reagents * The more drugs/hormone in the patient sample * The more drug --- to antibody (competing with bound Ab) * The more enzyme is available for converting the substrate to the product * A standard curve is used ---- the amount in the sample

- seperation - binds - quantify

--- is the interpretation of results and it includes :

- post analytical phase - includes: Validation of results Review of internal and external quality assurance results Internal – precision External - accuracy

interpretation of results :

1-Results are often reported as concentrations: Concentrations change when: -Amount of the analyte changes -Volume of solution changes e.g. with dilution (Beware differing units e.g. molarity vs. μg/ml) 2-Variability of results caused by: -Analytical factors and Biological factors 3-Uncertainty of Measurement - Estimates the difference between measured value and true value - -describes a level of confidence in the measurement 4-Factors affecting uncertainty - Sample collection and transport - Calibration, precision and bias of assay 5-Reporting uncertainty of measurement Plasma Sodium 141 mmol/L +/-0.64 mmol/L ( CHECK SLIDES FROM 52 PLSSS)

----- the range of values that represent 90 or 95% of reference population

reference range ( check slide 57)

Plasma alkaline phosphatase (total) activity Isoenzymes present in plasma derived from:-

Bone Liver Kidney Placenta 3rd trimester of pregnancy Intestine following fatty meal ( check slide from 59)

- selection of enzymes for diagnosis or prognostic purposes depends on:

1-Distribution of enzyme among various tissues -Tissue / plasma concentration gradient -intracellular localisation 2-Convenience of enzyme assay -Knowledge of plasma enzyme characteristics -Half-life in blood -Mode of clearance

why do we need to know all this - the lab results and reference ranges are affected by --- such as:

- labarotary methodology as: 1- substrate and substrae concnetration 2- buffer and pH 3- temp 5- presence of enzyme activator/inhibitor ( check slide 63 -62)