Clinical Chemistry part 2 Flashcards by Kamel Caragay

Organic molecules that are water insoluble, fat-soluble

Lipids and lipoproteins

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Building blocks of lipids; hydrocarbon chains with a terminal COO group

Fatty acids

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3 fatty acid molecules attached to one molecule of glycerol by Ester bonds; serves as main storage form of energy, insulator, shock absorber and integral part of cell membrane

Triglycerides

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Similar to triglycerides except that the third position on the glycerol backbone contains a phospholipid head group; contains polar and non-polar end; consitutent of cell membranes

Phospholipids

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Serves as part of cell membranes and as parent chain for cholesterol-based hormones

Cholesterol

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2 forms of cholesterol

Cholesterol esters and free cholesterol

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approximately 70% of total cholesterol

Cholesterol esters

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approximately 30% of total cholesterol

free cholesterol

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Typically spherical in shape with sizes ranging from 10 to 1200 nm

Lipoprotein

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Lipoproteins are composed of lipids and proteins called

apolipoproteins

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The size of lipoproteins particle correlates with its lipid content

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The various lipoproteins were separated through

Ultra centrifugation

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Located on the surface of lipoproteins particles; maintain structural integrity of lipoproteins; serve as ligands for cell receptors

Apolipoproteins

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Function of lipoproteins

Transport lipids

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Apo A-I location

HDL

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Apo A-II

HDL

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Apo A-IV

Chylos, VLDL, HDL

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Apo B-100

LDL, VLDL

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Apo B-48

Chylos

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Apo C-I

Chylos, VLDL, HDL

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Apo C-II

Chylos, VLDL, HDL

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Apo C-III

Chylos, VLDL, HDL

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Apo-E

VLDL, HDL

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Apo (a)

Lp(a)

Largest lipoprotein; least dense; highest TAG content; can block light: causes post-prandial turbidity

Chylomicrons

Transports exogenous or dietary TAG

Chylomicrons

2nd largest, 2nd least dense, 2nd highest TAG content

VLDL

Causes FASTING HYPERLIPIDEMIC TURBIDITY

VLDL

Transports endogenous or hepatic TAG

VLDL

Small; highest cholesterol content; transports cholesterol To peripheral tissues

LDLQ\

Target of cholesterol lowering therapy (statins)

LDL

Deposition of cholesterol

Atherosclerosis

Smallest, densest, highest protein content

HDL

Reverse transport of cholesterol

HDL

Triglycerides rich lipoproteins

CM, VLDL

Protein-rich lipoproteins

LDL, HDL

Highest TAG content

Highest cholesterol content

LDL

Highest protein content

HDL

Apo B containing

CM, VLDL, LDL

Alpha lipoprotein

HDL

Pre-beta lipoprotein

VLDL

Beta lipoprotein

LDL

Target for station therapy

LDL

Floating beta lipoprotein

Beta-VLDL

Increased in familial dysbetalipoproteinemia

Beta-VLDL

Sinking Pre-beta lipoprotein

Lp(a)

LDL-like particule; increased risk of premature coronary heart disease and stroke

Lp(a)

Seen in patients with biliary cirrhosis or cholestasis and in patients with mutations in the enzyme LCAT

LpX

Triglyceride content of CM

85-95%

Triglyceride content of VLDL

45-65%

cholesterol content of LDL

6-8%

Protein content of HDL

45-55%

Optimal LDL cholesterol Reference range

<100

Near optimal LDL cholesterol Reference range

100-129

Borderline high LDL cholesterol Reference range

130-159

High LDL cholesterol Reference range

160-189

Very high LDL cholesterol Reference range

>/= 190

Desirable total cholesterol Reference range

<200

Borderline high total cholesterol Reference range

200-239

High total cholesterol Reference range

>/= 240

High HDL cholesterol Reference range

>/= 60

Low HLD cholesterol Reference range

< 40

Normal TAG reference range

< 150

Borderline high TAG reference range

150-199

HIigh TAG reference range

200-499

Very high TAG reference range

>/= 500

Used to characterize lipid disorders

Fredrickson Classification

The Fredrickson Classification used ___ and ___ for CM to correlate clinical disease syndromes with laboratory phenotypes.

Electrophoresis and standing plasma test

Type 1 Hyperchylomicronemia (Familiar LPL deficieny)

Increased TG and CM

Type 2a (Familial Hypercholeseterolemia

Increased cholesterol and LDL

Type 2b (Familial Combined Hyperlipidemia)

Increased TG, Cholesterol, LDL, VLDL

Type 3 (Familial Dysbetalipoproteinemia)

Increased TG, Cholesterol, and VLDL

Type 4 (Familial Hypertriglyceridemia)

Increased TG, VLDL

Type 5

Increased all except LDL

Fasting requirement for Lipid profile

12 hours

2 parameters that can be measured nonfasting

TC and HDL-C

Prolonged tourniquet application can cause ___

Hemoconcentration

Reclined patients causes ___ values

Decreased

Preferred sample for lipid profile

Serum or plasma

Preferred sample in electrophoresis and ultracentrifugation (lipid profile)

Plasma

Capillary blood samples have generally __ values

Lower

Lipemic samples are seen when triglyceride levels exceed ___

4.6 mmol/L (400 mg/dl)

Cholesterol measurement (3)

1. Abell-Kendall method 2. Enzymatic methods 3. IDMS

Cholesterol measurement wherein the initial extraction with zeolite is used to remove sterols  redissolving of cholesterol  hydrolysis of cholesterol esters to cholesterol

Abell-Kendall method

3 components of the Liebermann-Burchard reagent

Glacial acetic acid, sulfuric acid, acetic anhydride

Absorbance of the Abell-Kendall method

410 nm

Reference method for Cholesterol measurement

GC-MS method

Gold standard for drug testing

GC-MS method

In the enzymatic method of cholesterol measurement, what is the first enzyme in the reaction

Cholesteryl esterase

Definitive method of cholesterol measurement

Isotope Dilution Mass Spectrometry (IDMS)

Triglyceride measurement (3)

1. Enzymatic method 2. Chemical methods 3. GC-MS method

In the enzymatic method of triglyceride measurement, what is the first enzyme in the reaction?

Bacterial lipase

Hydrolysis of glycerol is accomplished using alcoholic KOH  Oxidation of glycerol by periodic acid, forming formaldehyde and formic acid  formaldehyde combines with a variety of reagents forming colored products and fluorescence

Chemical methods of triglyceride measurement

Colorimetric method of triglyceride measurement

Van Handel and Zilversmit

Fluorometric method of triglyceride measurement

Hantzsch

Reagent in Van Handel and Zilversmit (TG)

Chromotropic acid

Product formed in Van Handel and Zilversmit

Blue colored compound

Reagent in Hantzsch method (TG)

Acetylacetone (diacetyl acetone; reactant of choice)

Produce formed in Hantzsch method

Strong absorption maximum at 412 nm and has a good fluorescence

New reference method for triglyceride measurements; involve the hydrolysis of fatty acids on triglycerides and the measurement of glycerol

GC-MS method

Range in density observed among lipoprotein classes is a function of lipid and protein content and enables fractionation by density using ___

Ultracentrifugation

Takes advantage of differences in size and charge

Electrophoresis

___ can be performed using unfractionated plasma or in plasma fractions that contain other serum proteins.

Electrophoresis

Most anodal lipoprotein

HDL

Lipoprotein electrophoretograms are usually visualized with a lipid-staining dye such as ___

Oil Red O Fat Red 7B Sudan Black B “OFS or Only Find Sun”

Depends on particle size, charge, and differences in the apolipoprotein content; primarily used in research labs only. It uses polyanions (heparin sulfate, dextran sulfate and phosphotungstate) and divalent cations such as magnesium, calcium and manganese)

Chemical precipitation

Uses antibodies specific to apolipoproteins to bind and separate lipoprotein classes

Immunoassays

Takes advantage of size differences in molecular sieving methods or composition in affinity methods

Chromatographic methods

Recall the Friedewald calculation

[Plasma TG/5] = mg/dl [Plasma TG/2.175] = mmol/l [Plasma TG/2.825] = gives more accurate estimate of VLDL-C , which is equivalent to [Plasma TG/6.5]

Involves large and medium sized arteries

Atherosclerosis

Increased risk of atherosclerosis

LDL

Decreased risk of atherosclerosis

HDL

Narrowing of blood vessels result in impaired blood flow and ischemia leading to Peripheral vascular disease Angina Ischemic bowel disease

Complications of atherosclerosis

Plaque rupture  thrombosis  ___

Myocardial infarction and stroke

Plaque rupture  embolization  ___

Atherosclerotic emboli

Weakening of blood vessel wall results in ___

Aneurysm

An extreme form of hypoalphalipoproteinemia (isolated decrease in circulating HDL)

Tangier Disease

Tangier disease is associated with HDL cholesterol concentrations as low as ____ in homozygotes, accompanied by total cholesterol concentrations of ___

1-2 mg/dl (0.03 -0.05 mmol/l; 50 – 80 mg/dl (1.3 – 2.1 mmol/L)

Tangier disease is associated with increased risk of premature ____

Coronary heart disease (CHD)

Linear polymers of amino acids

Proteins

Regulate metabolism Facilitate contraction in the muscle Provide structural framework Shuttle molecules in the bloodstream Component of the immune system

Functions of proteins

Number and types of amino acids in the specific amino acid sequence

Primary

Regularly repeating structures stabilized by hydrogen bonds between the amino acid within the protein (local folding; geometrically structured levels)

Secondary

Overall shape, or conformation of the protein molecule

Tertiary

Shape or structure that results from the interaction or more than one protein molecule, or protein subunits held together by noncovalent forces

Quaternary

Number and types of polypeptide units of oligometric proteins and their spatial arrangement

Quaternary

Most plasma proteins are synthesized in the ___ and secreted by the hepatocyte into the circulation

Liver

___ are synthesized in plasma cells

Immunoglobulins

___ are produced by endothelial cells and megakaryocytes

vWF

In liver failure, all proteins are decreased except

Immunoglobulin and VWF

The nitrogen content of serum protein is approximately ___

16%

Site of protein synthesis within the cell

Ribosomes

Indicator of malnutrition; binds thyroid hormones and retinol-binding protein

Prealbumin (Transthyretin)

Binds bilirubin, steroids, fatty acids; major contributor to oncotic pressure

Albumin

Protease inhibitor

Alpha-1-antitrypsin

Principal fetal protein

Aplha-1-fetoprotein

May be related to immune response

Alpha-1-acid glycoprotein

Binds to hemoglobin

Haptoglobin

Transports copper; peroxidase activity

Ceruloplasmin

Binds heme

Hemopexin

Immune response

Complement

Precursor of fibrin

Fibrinogen

Opsonin

C-reactive protein

Migrates before albumin in the serum protein electrophoresis

Prealbumin

Transport protein for thyroid hormones; transports Vitamin A by forming a complex with retinol-binding protein

Prealbumin (Transthyretin)

Decreased in hepatic damage, acute-phase inflammatory response and tissue necrosis

Prealbumin

Sensitive marker of poor nutritional status

Low prealbumin

Increased in patients receiving steroids, in alcoholism, and in chronic renal failure

Prealbumin

Negative Acute Phase Reactant (decreased in inflammation)

Prealbumin, Albumin, Transferrin

Protein present in the highest concentration in serum (2/3 of total protein)

Albumin

Provide nearly 80% of colloid osmotic pressure (COP) of intravascular fluid

Albumin

Buffers pH

Albumin

Binds to various substances in blood (hormones, drugs, electrolytes, unconjugated bilirubin)

Albumin

50% of calcium is transported by ___

Albumin

Decreased in liver disease, malnutrition, malabsorption, kidney loss (nephrotic syndrome), hemodilution Increased in hydration

Albumin

Most important function is the inhibition of the protease neutrophil elastase

Alpha-1-antitrypsin

Abnormal form of AAT can also accumulate in the liver and cause ___

Cirrhosis

Alpha-1-antitrypsin is a major component of a1-globulin band  deficiency of AAt seen as lack of an a1-globulin band on SPE

True

Excessive elastase breaks down elastin in alveoli leading to ___

Emphysema

Normal albumin-globulin ration

2:1

Hypoalbuminemia leads to

Edema (H20 goes out of the BV and enters the tissue)

One of the COPDs (chronic obstructive pulmonary diseases). Most common cause is smoking

Emphysema

Excessive inflammation or lack of AAT leads to destruction of alveolar air sacs  loss of elastic recoil and collapse of airways during exhalation  obstruction and air trapping

Emphysema

“Pink puffers” “barrel-chest”, hypoxemia

Emphysema

Synthesized by the developing embryo and fetus; thought to protect the fetus from immunologic attack by the mother. No known function in normal adults

Alpha-1-fetoprotein

Elevated AFP

Neural tube defects, presence of twins

Low AFP

Increased risk for Down syndrome

Tumor marker for hepatocellular carcinoma, some testicular carcinomas

Alpha-1-fetoprotein

Cooper-containing (contains >90% of total serum copper)

Ceruloplasmin

Used in diagnosis of Wilson’s disease

Ceruloplasmin (decreased)

Decreased levels of ceruloplasmin. Excess storage of copper in various organs. Liver  hepatic cirrhosis Brain  neurologic damage Cornea  Kayser-Fleischer rings

Wilson’s disease

Total serum copper decreased Free serum copper increased Urinary copper increased

Wilson’s disease

Large protein that inhibits proteases such as trypsin, thrombin, kallikrein, and plasmin

Alpha-2-macroglobulin

Increased in nephrotic syndrome (large size aids in its retention)

Alpha-2-macroglobulin

Glomerular disorder characterized by proteinuria (>3.5 g/day) Disruption of the electrical charges that produce the tightly fitting podocyte barrier resulting in massive loss of protein and lipids

Nephrotic syndrome

Manifestations of nephrotic syndrome

Hypoalbuminemia – pitting edemia Hypogammaglobulinemia – increased risk of infection Hypercoagulable state – due to loss of anti-thrombin III Hyperlipidemia and hypercholesterolemia – may result in fatty casts in the urine

Marker for intravascular hemolysis

Haptoglobin

Bind free hemoglobin to prevent loss of hemoglobin and its constituent iron into the urine

Haptoglobin

Haptoglobin-hemoglobin complex is removed by

Reticuloendothelial system (mainly in the spleen)

Transports two molecules of ferric iron

Transferrin

Major component of the beta-globulin fraction

Transferrin

Tested to determine cause of anemia

Transferrin

Scavenge heme released or lost by the turnover of heme proteins such as hemoglobin  protect body from oxidative damage that free heme can cause

Hemopexin

Diagnostic of hemolytic anemia

Low hemopexin levels

Precipitates with C substance, a polysaccharide of pneumococci

C-Reactive Protein

Functions in opsonization

C-Reactive Protein

One of the first acute-phase proteins to rise in inflammatory disease

C-Reactive Protein

High or increasing amount of CRP suggests an ___

Acute infection or inflammation

___ is the same protein but is named for the newer, monoclonal antibody-based test methodologies that can detect CRP at levels below 1 mg/L

High-sensitivity CRP

The ___ test determines risk of cardiovascular disease

High-sensitivity CRP

Glycoprotein produced by fetal membranes responsible for the cellular adhesiveness of placenta and membranes to the decidua

Fibronectin

Fetal ___ is produced at the boundary between the amniotic sac and the decidua (the lining of the uterus) and functions to maintain the adherence of the placenta to the uterus

Fibronectin

Fibronectin test for assessment of the risk for ___ in women between 24 to 35 weeks gestational age

Preterm delivery

Proteolytic fragments of collagen I formed during bone resorption

Cross-Linked C-Telopeptides

Biochemical marker of bone resorption that can be detected in serum and urine

Cross-linked C-telopeptides

Gold standard for the detection of myocardial infarction

Troponin

Govern excitation-contraction coupling in muscle

Troponin

Used as an AMI indicator because of specificity and early rise in serum concentration

Troponin T or Troponin I

Troponin T rises within ___ Peaks in ___ Returns to normal in ___

3-4 hours 10-24 hours 10-24 days

Troponin I rises within ____ Peaks in ___ Returns to normal in ____

3-6 hours 14-20 hours 5-10 days

Earliest protein marker for Myocardial infarction

Myoglobin

Heme-containing protein that binds oxygen with cardiac and skeletal muscle. Levels are related to muscle mass and activity (reasonable sensitivity but poor specificity)

Myoglobin

Increased in skeletal muscle injuries, muscular dystrophy, and AMI

Myoglobin

Myoglobin rises in ___ Peaks in ___ Returns to normal in ___

1-3 hours 5-12 hours 18-30 hours

Myoglobin is not tissue specific. It is better used as a negative predictor in the first 2-4 hours following chest pain

True

Neurohormones that affect body fluid homeostasis (through natriuresis and diuresis) and blood pressure

Natriuretic peptides

Found in largest concentration in the left ventricular myocardium but are also detectable in atrial tissue as well as in the myocardium of the right ventricle

NT-proBNP and BNP

BNP has become a popular marker for ___

Congestive Heart failure

A1- globulins

A1-antitrypsin A1-fetoprotein A1-acid glycoprotein A1-lipoprotein A1-antichymotrypsin Inter-a-trypsin inhibitor Gc globulin

A2 globulins

Haptoglobin Ceruloplasmin A2-macroglobulin “CHA”

Beta globulins

Pre-b-lipoprotein b-lipoprotein Transferrin Hemopexin B2-microglobulin Complement Fibrinogen CRP

y-Globulins

IgA IgD IgE IgG IgM CRP

Share the property of showing elevations in concentrations in response to stressful or inflammatory states that occur with infection, injury, surgery, trauma, or other tissue necrosis.

Acute phase reactants

CRP response time Normal concentration (mg/dl) Increase Function

6-10 hours (preferred marker for inflammation) 0.5 1000x Opsonization, complement activation

Serum amyloid A response time Normal concentration (mg/dl) Increase Function

3.0 1000x Removal of cholesterol

A-1-antitrypsin response time Normal concentration (mg/dl) Increase Function

24 hours 200-400 2-5x Protease inhibitor

Fibrinogen response time Normal concentration (mg/dl) Increase Function

24 hours 110-400 2-5x Clot formation

Haptoglobin response time Normal concentration (mg/dl) Increase Function

24 hours 40-200 2-10x Binds hemoglobin

Ceruloplasmin response time Normal concentration (mg/dl) Increase Function

48-72 hours 20-40 2x Binds coppere, oxidizes iron

C3 response time Normal concentration (mg/dl) Increase Function

48-72 hours 60-140 2x Opsonization, lysis

Mannose-binding protein response time Normal concentration (mg/dl) Increase Function

? 0.15-1.0 ? Complement activation

Total protein level less than the reference interval

Hypoproteinemia

Occurs in any condition where a negative nitrogen balance exists (excessive loss, decreased intake, decreased synthesis, accelerated catabolism)

Hypoproteinemia

Increased in total plasma proteins

Hyperproteinemia

Not an actual disease state, but is the result of dehydration

Hyperproteinemia

Abnormal proliferation of plasma cells  produce many Ig (which are abnormal)

Multiple myeloma

Total Protein analysis (4 methods)

Kjeldahl Refractometry Biuret Dye binding

Digestion of protein; measurement of nitrogen content Reference method of total protein analysis; assume average nitrogen content of 16%

Kjeldahl

Conversion factor in Kjeldahl method

6.25 or 6.54

Measurement of refractive index due to solutes in serum Rapid and simple; assume nonprotein solids are present in same concentration as in the calibrating serum

Refractometry

Formation of violet-colored chelate between Cuprous ions and peptide bonds Routine method; requires at least 2 peptide bonds and an alkaline medium

Biuret

Protein binds to dye and causes a spectral shift in the absorbance maximum of the dye For research use

Dye binding

Globulins are precipitated in high salt concentrations; albumin in supernatant is quantitated by biuret reaction Labor intensive

Salt-precipitation (Historical method)

Albumin binds to dye; causes shift in absorption maximum; Nonspecific for albumin

Methyl orange

Albumin binds to dye; causes shift in absorption maximum; Many interferences (salicylates, bilirubin)

HABA

Albumin binds to dye; causes shift in absorption maximum; Sensitive; overestimates low albumin levels; most commonly used dye

Bromcresol green

Albumin binds to dye; causes shift in absorption maximum; Specific, sensitive, precise

Bromcresol purple

Protein separated based on electric charge; accurate, gives overview of relative changes in different protein fractions

Electrophoresis

Performed when an abnormality in the total protein or albumin is found. Separation of proteins based on their charge density

Protein electrophoresis

Regions in protein electrophoresis are stained using:

Coomassie Blue, Amido Black, Ponceau S

Beta-gamma bridging electrophoretic pattern

Cirrhosis

Monoclonal spike electrophoretic pattern

Multiple myeloma

Increased a2, increased b2, decreased albumin electrophoretic pattern

Nephrotic syndrome

Decreased a1-antitrypsin electrophoretic pattern

Emphysema

Increased beta region electrophoretic pattern

Use of plasma instead of serum (fibrinogen)

The most significant finding from an electrophoretic pattern is ___

Monoclonal immunoglobulin disease

Non-protein nitrogen compounds

Urea Amino acids Uric acid Creatinine Creatine Ammonia

NPN present in the highest concentration in blood

Urea

Major excretory product of protein metabolism

Urea

Concentration of urea in the plasma is determined by

Renal function and perfusion, Protein content of the diet Rate of protein catabolism

Most common cause of death or liver failure

Increased ammonia

Evaluate renal function Assess hydration status Aid in diagnosis of renal disease Verify frequency of dialysis

Clinical applications of urea

Urea nitrogen concentration is converted to urea concentration by multiplying

2.14

Most used method in determining urea concentration

Enzymatic methods

Proposed reference method for the urea determination

IDMS

Elevated concentration of urea in the blood

Azotemia

Very high plasma urea concentration accompanied by renal failure is called

Uremia or the uremic syndrome

Prerenal azotemia

Congestive heart failure Shock, hemorrhage Dehydration Increased protein catabolism High-protein diet

Renal azotemia

Acute and chronic renal failure Renal disease (glomerular nephritis, tubular necrosis)

Postrenal azotemia

Urinary tract obstruction

Decreased urea concentration

Low protein intake Severe vomiting and diarrhea Liver disease Pregnancy

Product of catabolism of purines (guanine and adenosine)

Uric acid

Filtered by the glomerulus and secreted by the distal tubules into the urine, but mostly reabsorbed in the proximal tubules and reused

Uric acid

Relatively insoluble in plasma, and a high concentrations, can be deposited in the joints and tissues, causing painful inflammation

Uric acid

Uric acid is mostly present as ___ in plasma, wherein it is insoluble at around pH 7

Monosodium urate

Increased uric acid in urine “Orange sand” in diapers UA crystals

Lesch-Nyhan syndrome

Uric acid determination (phosphotungstic acid and tungsten blue)

Caraway method

Proposed reference method for uric acid

IDMS

Uric acid determination or method where there are problems with turbidity and several common drugs interference

Colorimetric

Preferred marker for kidney function (glomerular filtration)

Creatinine or creatine

Creatinine is formed from ___ (synthesized primarily in the liver from arginine, glycine, and methionine) and creatine phosphate in muscle

Creatine

Excreted in plasma at a constant rate related to muscle mass

Creatinine

Daily excretion is fairly stable thus it is commonly used to assess renal filtration function

Creatinine

Determine sufficiency of kidney function and severity of disease Monitor the progression of kidney disease Measure of completeness of 24 hour collections

Creatinine

Increased creatinine concentration

Abnormal renal function Muscle disease

Normal BUN/Creatinine ratio

10:1 – 20:1

The BUN/Creatinine ratio rises in prerenal disease to

>20:1

In true renal disease, both BUN and creatinine rise together, maintaining BUN/Creatinine ratio at __

10-20:1

__ performed directly on sample; detection of color formation times to avoid interference of noncreatinine chromogens

Jaffe-kinetic

Creatinine in protein-free filtrate adsorbed onto Fuller’s earth (aluminum magnesium silicate); then reacted with alkaline picrate to form colored complex Lloyd’s reagent (sodium aluminum silicate)

Jaffe with adsorbent

Highly specific, accepted reference method for creatinine

IDMS

Formed through the deamination of amino acids during protein metabolism

Ammonia

Ammonia is removed from the circulation and converted to __ in the liver

Urea

Provide useful information on clinical conditions such as hepatic failure, Reye’s syndrome and inherited deficiencies of the urea cycle enzymes

Ammonia

Indicator in the spectrophotometric method of ammonia determination

Bromphenol blue

Enzymatic method of ammonia determination

Glutamate dehydrogenase

Specimen consideration for ammonia

Should be iced, Avoid smoking prior to collection

Increased BUN, Normal creatinine, Increased BUN/Crea ratio

Prerenal azotemia

Increased BUN, increased creatinine, Normal Ratio

Renal, postrenal azotemia

Largest and heaviest internal organ

Liver

Weight of the liver

1.2 – 1.5 kg

Ligament the divides the right and left lobe of the liver

Falciform ligament

Functional units of the liver

Lobules/ acini

Supplies oxygen-rick blood in the liver (main blood vessel)

Hepatic artery

Supplies deoxygenated blood to the liver; delivers blood from the gallbladder, spleen, GIT to the liver

Hepatic portal vein

Capillary-like blood vessel where blood from hepatic artery and hepatic vein miX

Sinusoids

Small passages in the liver

Canaliculi

Perform most of the liver function; contribute to the regenerative properties of the liver

Hepatocytes

Derived from macrophages; found in sinusoids

Kupffer cells

Synthesize nitric oxide (regulate blood flow)

Stellate/Ito cells

Liver stem cells (regeneration of hepatocytes and bile ducts)

Oval cells

Carries bile

Common bile duct

Layer of loose connective tissue in the liver

Glisson’s capsule

Covers the exterior portion of the liver

Peritoneum

Synthetic function of the liver

Produce substances such as proteins, bile salts, lipids, and carbohydrates

Storage function of the liver

Glycogen, iron, amino acids, and some lipids

Excretes bilirubin, bile acids, and ammonia

Detoxification function of the liver and drug metabolism

Convert toxic substances

Bile acids are conjugated with amino acids (taurine, glycine) to form

Bile salts

Aids in absorption and digestion of vitamins

Bile salts

Major metabolite of heme; orange-yellow pigment derived from hemoglobin

Bilirubin

Daily bilirubin production

250-300 mg

Bilirubin is mainly transported by __

Albumin

Bilirubin production is contributed by __

85% liver, spleen, BM (reticuloendothelial) 15% ineffective erythropoiesis, heme-containing proteins/hemoproteins

3 heme-containing proteins/hemoproteins

Cytochromes Peroxidases Myoglobin

How many grams of albumin is produced a day by the liver

12 g

One of the synthetic functions of the liver is the metabolism of cholesterol into ___

Primary bile acids

Promote fat absorption and emulsification

Primary bile acids

2 types of primary bile acids

Deoxycholic acid and chenodeoxxycholic acid

Unconjugated, nonpolar, water insoluble, indirect bilirubin, hemobilirubin

Conjugated bilirubin, polar, water soluble, direct bilirubin, cholebilirubin

First type of bilirubin formed

Type of bilirubin not normally present in the urine

Type of bilirubin that slowly reacts with diazo reagent

Bilirubin that is normally in the circulation

Type of bilirubin that needs an accelerator or solubilizer

Bilirubin that is freely filtered in plasma and present in the urine

Bilirubin that directly reacts with diazo reagent

Aka biliprotein; a bilirubin that is covalently bound to albumin thus contributes to the direct bilirubin

Delta bilirubin

Formula for total bilirubin

Total bilirubin = unconjugated + conjugated + delta bilirubin

“Jaune” = yellow Yellow discoloration of the skin, eyes, and mucous membranes due to bilirubin retention

Jaundice

Serum bilirubin in adults and neonates

2-3 mg/dl (adults) 5 mg/dl (neonates)

Amount of bilirubin that becomes noticeable to the naked eye

> 3 mg/dl

Unconjugated hyperbilirubinemia; Mild type of jaundice that occurs prior to liver metabolism

Pre-hepatic jaundice

Most common cause of pre-hepatic jaundice

Hemolytic anemia Ineffective erythropoiesis

Results from intrinsic liver disease due to defects in bilirubin metabolism and transport; due to diseases resulting to hepatocellular injury

Hepatic jaundice

Inherited, autosomal recessive mild form of unconjugated hyperbilirubinemia Serum bilirubin: 1.5 – 3 mg/dl The gene responsible for the expression of the enzyme UDPGT is mutated or due to transport deficit across the hepatocyte membrane

Gilbert syndrome

Problem in conjugation deficit

Criggler-Najjar syndrome

UDPGT absent; serum bilirubin >20 mg/dl; kernicterus; therapy is liver transplant

Type 1 Crigler-Najjar syndrome

Partial UDPGT deficiency (25% activity); serum bilirubin 5-20 mg/dl; no kernicterus; patients responds to UV therapy

Type 2 Crigler-Najjar syndrome

Transporter gene deficit

Dubin-Johnson syndrome

Predominantly elevated conjugated bilirubin; excretion of bilirubin is impaired; liver biopsy shows dark brown pigment in hepatocytes due to accumulation of lipofuscin-like pigment; B2 cannot be transported out of the hepatocyte

Dubin-Johnson syndrome

Conjugated hyperbilirubinemia similar to Dubin-Johnson syndrome but without liver pigmentation

Rotor syndrome

Transient familiar neonatal hyperbilirubinemia; presence of UDPGT inhibitor (antibody); mild hyperbilirubinemia that lasts for the first 2-3 weeks of life; increased B2

Lucey-Driscoll syndrome

3 types of unconjugated hyperbilirubinemia in the jaundice of the newborn

Physiologic jaundice of the newborn Hemolytic disease (HDFN) Breastmilk hyperbilirubinemia

In breastmilk hyperbilirubinemia what inhibits the conjugation

Alpha glucoronidase

Conjugated bilirubin is >1.5 mg/dl; most important causes are idiopathic neonatal hepatitis and biliary atresia

Conjugated hyperbilirubinemia in the jaundice of the newborn

Results from biliary obstructive disease

Post hepatic jaundice

Most common causes of post hepatic jaundice

Gall stones or tumors in the biliary tree

Common cause of gallstones

Ketodiets (intermittent fasting) High fat diet and steatorrhea

Increased serum B1, Normal serum B2, increased serum total bilirubin

Pre-hepatic jaundice

Increased serum B1, decreased serum B2, increased serum total bilirubin

Gilbert syndrome, Crigler-Najjar syndrome, jaundice of the newborn, Lucey-Driscoll syndrome

Normal serum B1, increased serum B2, Increased serum total bilirubin

Dubin-Johnson syndrome, Rotor syndrome, Post-hepatic jaundice

Viral hepatitis that causes hepatocellular injury

Hepatitis A,B,C,D,E,G

Acute injury of hepatocytes caused by toxic drugs, ischemia, or immunologically mediated injury

Acute hepatitis

Accumulation of fats as a consequence of insulin resistance

Non-alcoholic fatty liver disease and non-alcoholic steatohepatitis

Chronic liver inflammation that persists for at least 6 months

Chronic hepatitis

Diffused fibrosis with nodular regeneration (end stage of scar formation)

Liver cirrhosis

Causes mild hyperbilirubinemia with three fold rise in transaminases (AST and ALT); has a strong association with intake of aspirin; characterized by non-inflammatory encephalopathy and fatty degeneration of the liver; usually affects children 2-14 years old after a viral infection – flu and chickenpox (most harmful in affecting brain and liver)

Reye Syndrome

Methods of bilirubin determination

Ehrlich reaction Van den Berg reaction Jendrassik-Grof method Evelyn-Malloy method Bilirubinometry

Bilirubin + diazotized sulfanilic acid  azobilirubin Initially done on urine and stool samples (freshly collected 24 hour samples)

Ehrlich reaction

Bilirubin + diazotized sulfanilic acid  azobilirubin Done using serum samples; Diazotized sulfanilic acid is formed by reacting sulfanilic acid with sodium nitrite and hydrochloric acid

Van den Berg reaction (and Muller reaction)

Results of the Van den Berg reaction at neural pH and at high or low pH

At neutral pH  reddish purple At low or high pH  blue

Diazotized sulfanilic acid + accelerator/solubilizer (caffeine sodium benzoate) ; Buffer: sodium acetate Strong alkaline tartrate is added to convert original purple color into blue (measured spectrophotometrically at 600nm)

Jendrassik-Grof method

Advantages of Jendrassik Grof method: Insensitive to sample pH changes and variation in protein concentration of sample Not affected by hemoglobin up to 750 mg/dl Has adequate optical sensitivity even for low bilirubin concentrations

True

Accelerator in Evelyn-Malloy method

50% methanol

Azobilirubin: red to reddish purple color in acid pH (measured at 560 nm)

Evelyn-Malloy method

Done in neonates Transcutaneous bilirubin concentration device; Measurement of reflected light from the skin using 2 wavelengths, providing a numerical index based on spectral reflectance; Has a positive interference with carotenoid compounds thus cannot be done in adults

Bilirubinometry

Diazo reagents

Diazo A: 0.1% Sulfanilic acid + HCl Diazo B: 0.5% Sodium nitrite Diazo Blank: 1.5% HCl

Tests Measuring Hepatic Synthetic Ability

Total Protein determination Prothrombin time – Vitamin K response test Test for albumin Albumin/Globulin ratio

Total bilirubin

Up to 1.0 mg/dl

Direct bilirubin

<0.5 mg/dl (other references 0 – 0.2 mg/dl)

Indirect bilirubin

0.2 – 0.8 mg/dl (other references 0 – 1 mg/dl)

Conversion factor of bilirubin

17.1

Critical value of bilirubin

>18 mg/dl (may cause kernicterus)

Intamuscular injection of 10 mg Vitamin K everyday for 1 – 3 days

Vitamin K response test

Results in vitamin K response test

Prolonged: Intrahepatic disorder Normal: Extrahepatic disorder Consistently prolonged: Loss of hepatic synthetic ability of liver

Tests measuring conjugation and excretion function

Bilirubin assay Bromosulfophthalein dye exretion test Urobilinogen test

2 methods under Bromosulfophthalein dye exretion test

Rosenthal White method MacDonald

Rosenthal White Method

Double collection method After 5 mins: 50% dye retention After 30 mins: 0%

MacDonald (Reference method)

Single collection method NV: After 45 mins +/- 5% retention of dye

Ehrlich’s reagent

p-dimethyl aminobenzaldehyde

Uses Ehrlich’s reagent Specimen: 2-hour freshly collected urine or stool sample Positive reaction: red color production

Urobilinogen Test

Consists of binding site and catalytic site; a site where substrate interacts with the enzyme

Active site

Site other than the active site; binds to regulator molecules

Allosteric site

Non-protein substances needed for maximal activity of the enzyme

Cofactors

Inorganic cofactors

Activators (anions and cations)

Organic cofactors

Coenzymes (acts as co-substrate in enzyme reactions)

Complete active enzyme system with full catalytic activity; apoenzyme + prosthetic group

Holoenzyme

Inactive form of an enzyme

Proenzyme/ Zymogen

Multiple forms of serum proteins that are functionally related; results from post translational modifications

Isoform

Multiple forms of an enzyme that catalyzes the same biochemical reaction

Isoenzyme

6 classes of enzymes

Oxidoreductases Transferases Hydrolases Lyases Isomerases Ligases

Catalyzes an electron transfer or oxidation-reduction reaction between 2 substrates

Oxidoreductasses

Catalyzes the transfer of various chemical groups from one molecule to another

Transferases

Catalyzes the hydrolysis of various bonds with the addition of water; involves splitting of molecules

Hydrolases

Catalyzes the removal of groups from substrates without hydrolysis; formation of double bonds

Lyases

Catalyzes the interconversion of geometric, optical or positional isomers within a molecule

Isomerases

Catalyzes the joining of two substrate molecules coupled with breaking of the pyrophosphate bond in ATP

Ligases (synthetases)

Rate of reaction is almost directly proportional to substrate concentration at a low level

First order kinetics

Substrate concentration is high enough to saturate all available active sites of the enzymes; Reaction is independent of substrate concentration Reaction rate depends only on enzyme concentration

Zero-order kinetics

Enzymes are affected by changes in ___ as it is a factor in their stability

Most favorable pH value at which enzyme is most active

Optimum pH (7.0 – 8.0)

ALP optimum pH

9.0 – 10.0

ACP optimum pH

5.0

pH are controlled using ___

Buffer solutions

Optimum temperature for most enzymes is ___

37 deg C

Increased temperature = increased rate of ___

Chemical reaction

For every ___ increase in temperature, rate of reaction is doubled until it is denatured

10 deg C

Absence means enzyme activity may not occur. In excess, may inhibit enzyme activity

Cofactors

Substrate competes with active site

Competitive inhibitor

Substrate competes other than the active site

Non-competitive inhibitor

Substrate competes with the substrate-enzyme complex

Uncompetitive inhibitor

Enzyme concentration is measured in fixed period of time; the reaction is stopped and a measurement is made

Fixed time assay (End point assay)

Multiple measurements (absorbance change) are made at specific time interval

Continuous monitoring assay

Amount of enzyme that will catalyze the reaction of 1 umol of substrate in 1 minute

1 International Unit

Amount of enzyme that would catalyze the reaction of 1 mole substrate in one second

1 Katal

EC numerical code of AST

2.6.1.1

Tissue source of AST

Cardiac tissue, liver, skeletal muscle

AST is previously known as

SGOT

Aspartate + alpha-ketoglutarate ____ + ____

Oxaloacetate, glutamate

Coenzyme of AST

Pyridoxal phosphate

2 enzyme fractions of AST

Cytoplasmic AST Mitochondrial AST

Most predominant AST in the serum

Cytoplasmic AST

Mitochondrial AST becomes highest than cytoplasmic AST in serum in ____

Cellular necrosis

Normal  cAST > mAST Necrosis 

cAST < mAST

AST highest elevation: 5 or more x ULN

Acute hepatocellular disorder Myocardial infarction Circulatory collapse Acute pancreatitis Infectious mononucleosis

AST 3 or more x ULN

Stop the medication (potential hepatotoxic drug)

AST in AMI

Rise within 6-8 hours Peaks at 24 hours Returns to normal within 3-5 days

Marked elevation of AST is observed in __

Viral hepatitis and AMI

Hemolysis increases AST concentration (T/F)

AST activity is stable in serum for ___ at refrigerated temperature

3-4 days

Methodologies for AST

Karmen method Reitman-Frankel method Diazonium salt reaction

Coupled enzymatic reaction Basis for IFCC Malate dehydrogenase is added: oxaloacetate + NADH  Malate + NAD

Karmen Method (AST)

In the Karmen method, the rate of decrease in absorbance at __ due to the formation of NAD is directly proportional to the activity of AST in the sample

340 nm

Optimal pH of Karmen method for AST determination

pH 7.3 – 7.8 (7.5)

Colorimetric method of AST determination Substrate: aspartic alpha-ketohlutarate Color developer: 2,4-dinitrophenylhydrazone

Reitman-Frankel method

Final reaction of Reitman-Frankel method of AST determination

Blue-colored complex, measured at 505 nm

The Reitman-Frankel method lacks specificity because it reacts with any ___

Keto-compound

Colorimetric method of AST determination Stabilized diazonium salt + oxaloacetic acid  red colored compound Formation of diazonium derivatives

Diazonium salt reaction

EC numerical code for ALT

2.6.1.2

Tissue sources of ALT

Highest concentration in liver and kidney, smaller amount in cardiac tissue and skeletal muscle

ALT is formerly known as

SGPT

More liver specific than AST

ALT (ALT>AST)

Involved in the reversible transfer of an amino group between alanine and alpha-ketoglutarate Alanine + alpha-ketoglutarate  ___+ _____

Pyruvate; glutamate

ALT has the highest elevation in __

Acute viral hepatitis

De Ritis ration (AST/ALT quotient) >1

Liver cirrhosis, metastatic carcinoma

De Ritis ration (AST/ALT quotient) 3-4:1

Alcohol-induced liver disease

ALT is stable in serum for ___ days at 4 deg C

3-4

ALT is unaffected by hemolysis

True

In liver disease, ALT > AST, except in __

Hepatic cirrhosis and liver neoplasia

ALT is increased, HCV (-), accept or reject donor?

Defer (consider window period)

Acute hepatocellular injury (AST/ALT ratio)

AST>ALT

Methodologies for ALT determination (4)

Wroblewski and La Due Reiman-Frankel method Diazonium salt reaction

Coupled enzymatic reaction (ALT determination) Basis for the IFCC recommended method Lactate dehydrogenase is added to convert endogenous pyruvate in the sample into lactate (excess pyruvate is inhibitory to ALT activity) Pyruvate + NADH  lactate + NAD (340 nm)

Wroblewski and La Due

Optimal pH in Wroblewski and La Due (ALT determination)

pH 7.3 – 7.8 (7.5)

Substrate in Reiman-Frankel method (ALT determination)

Alpha-ketoglutarate

Anticoagulant of choice for TDM

Heparin

Clinical Chemistry part 2 Flashcards

(463 cards)