Clinical Chemistry 431 PDF

 Lecture Le discussion Lecture 1 Case Study (multiple myeloma...

 Lecture Le discussion Lecture 1 Case Study (multiple myeloma 37-8 1) biuret method Cu2 alkaline soln. Presenting: March 6 , fig , containg in an sample would nu nemoglobin & 2) hemolysis: changes color electro : deviation hep ↓ N lower &2 Each BU junction class higher 1) Presentations 3) 2) Lecture Lecture 6 : Lecture S 3) mini quiz Liver monoclonal HMG-COA reductase , Quiz Protein most proteins are synthesized : Alipoprotein by the hepatic parenchymal cells A paraprotein is : monoclonal immune Lecture 3 : Lecture 1 mini quiz Enzyme assay single measurement Fixed time assay International Unit of enzyme activity 1 micromole of substrate per minute Lecture 4: Lecture y mini quiz Higest CK isoenzymes of healthy Ck MM - Children have ALP activity higher ALP leaks Osteoblasts during from normal growth Lecture 5 : Lecture mini 4 quiz Loss of in arterial blood supply oxugen Ischemia Condition from ineffective blood pumping of Congested heart failure Lecture 6 : Lecture 5 mini quiz Oxygen binding protein of Cardiac + Skeletal myoglobin Creatine Kinase isoenzymes "Cardia isoenzyme CK - 2  Exam 1 study Substrate concentration · Cryoglobulin (up c) : · Proteins 35 that precipitate at lower temps to · 3 types Type 1 : associated multiple myeloma protein (25 %) - wi [S] - Type 2: mixed multiple myeloma protein w/ other proteins (25 % ) substrate concentration · - Type 3 : mixed polyclonal proteins 1st order Kinetics: rate is linearly dependent on amount of substrate · Type associated - 2 + 3: often / immune complex disease ① order Kinetics: rate is independent on substrate concentration · · Clinical assesment Saturation of enzyme active sites Refrigerate - samples 12 hr O Limited # of active sites ; at low conc of substrate rate of rxn will be dep. On resuspend precipitated protein in warm Saline - Substrate Conc b) there will be effective unfilled. a large conc of active characterize by Serum protein electrophoresis (SPE) , - Sites immunofixation electrophoresis (IFE) · Limited # of active sites , enzyme is , constant as substrate increases, more active Sites filled , until saturated it the rxn rate is indep Of are making appear as bands in CSF Oligoclonal substrate conc. · Detection of in CSF of mS Ig patients · Observed in 90-95 % of ms patients Inhibitors · Indicative of synthesis Win the CNS · Competitive: bind to active site Ig · Super dark blue bands · non competitive: bind enzyme at site other than active · uncompetitive : bind to ES complex Serum Protein Electrophoresis : Sequence of mobility o Creatine Kinase (CPK , CK) Proteins Separated based on charge Assay · Proteins are amphoteric : can be (t) or () based on put Creatine > Phosphocreatine J ATP ADD · Isoelectric point : pl protein has net C where charge Transfer phosphate group - c) muscle contraction pH > isoelectric point : protein has net charge ↳ C+ terminal · 2 subunits (M B) migrate towards anode + - pH) isoelectric point : protein has net (t) charge · Separate chromosomes (19 + 14) ↳ C-terminal) · Found i n Cytosol of stricted muscle migrate towards cathode , t brain hear , · Speed of migration depends largely on the degree of ionization o f 3 primary isoenzymes buffer system (1) - pH of CK-BB brain the protein at the the : predominant in electric field of molecule CK-MB(z) : - Also depends size mainly in cardial muscle types of muscle on strength , shape , , also other cathode anode temp , + buffer characteristics - T -CK-MM(y) : predominant in Skeletal muscle + cardiac tissue I Can be · done on urine or CSF sample I measured by Coupled rxns , I · concentrated sensitive I Serum preferred (heparin EDTA , citrate) staining must be or , I I methods used; identification of bence-jones  · Creatine Kinase Assay cont. Elevated in hear t disease · other isoenzymes myocardial infarction - ↳ Ck (type macro) CK-2 CK-3 - - M+ 2 , in others ↳ · Mitochondrial isoenzyme Childbirth & located between inner +outer membranes of mitoch. - increased CK-1 ↳ indicator of severe illness OReyes Syndrome - (type Ig) Total CK increase - Macro-Ck 1 , complex CK between CK-BB + ↳ conjugate to get bigger Assay can wh Ig O ↑ origin Change in Abs 340 measured (NADINADH) I - ( +) Conjective Heart Failure 2 · Edema rest of body in lungs , · macro-Ck caused by coronary artery disease , cardiomyopathies , myocarditis · CK-3 valvular disease, Type 1 : most often CK-1 complexed w/ IgG ; sometimes WI IgA cardic arrhythmias -Associated w/ gastrointestinal disorders , adenomal carcinoma , myocardial/vascular Inability of heart to pump blood effectively ~ Associated w/ increased lower cardia output - mortality · Left side fails: fluid accumulates in lungs - Often in women > Sours - · Type 2 : predominantly in adults severely ill wh malignancy or liver disease, pulmonary edema myocardial disease -reduced output children w/ to systemic circulation Inter fere wi assay of CK-2 byion-exchange or immunoinhibition -Kidneys retain fluid · Differentiate from type I by fails : fluid accumulates in systemic venous higher activation Right circulation - side energy · from Both types are heat stable t distinguishable other CKisoenzymes -general edema poor blood - CK isoenzymes to lungs · Clinical decreased Significance output to arterial circulation Diagnosis · Elevated in muscle disorders , before symptoms · muscular dystrophy Cesp Duchemes. o Blood urea nitrogen (BUN) (MM) nitrogen - - Usually CK-3 measure serum urea - -CK-2 found in cases of diseased/damaged Skeletal muscle enzymatic (indirect : crease + ammonia) , direct (Fearon rxn) failure - : Elevated in CNS disease Increased assoc. wi renal - CHF ↑ BUN - CK-3 : acute disease , Cerebral ischemia, intervention w i normal creatinine Cerebrovascular neurosurgical : - · CK-1 : head trauma Uremia : increased blood ure a · creatinine , n2-containing · Thyroid disease Azotemia : increase in end products, area , Elevated hypothyroidism (5-50x cramping( in , total : due to muscle and uric acid ↳ mostly CK-b , some CK-2 OBNP Decreased in hyperthyroidism 400pg/mL : unlikely 100 : CHE 40049/mL likely - 74009g/mL : CHF very likely Enzymatic markers of M I Brain Natriuretic Peptide (BNP) · Total CK Released in response to ventricle vol. expansion + pressure overload O CK-1 (BB) : predominantly in brain works in tandem wi atrial natriuretic protein CK-2 (MB) : relatively heart Diagnosis of CHE · specific to Mar t ; 10-20 % of total (Kin < 2% total CK in skeletal Muscle 100pg/mL : CHF unlikely · [K-3 (MM) : cardiac + Skeletal 100 - 400 pglmc: CHF likely < Ck - 2 400pg/mL : CHF very likely · · Elevated 4-6 hr post MI , peaks ~24hr , normal 48-72 hr Provides prognostic info · must be serially measured O-3 , 3-6 , 6-9 , 12-24 hr after presentation o measured kM constant Cenzymes] · -Electrophoretically Michaelis-menten constant [substrate) - Enzymatically when V Vmax/2 - = (k -z = (total (k) - [(total m) + (total BT] -constant for a given enzyme under given conditions mass assays ↳ immuno / Ab specific for CK Lipoproteins " limit : 1 pg/L Chylomicrons ↑ · detects early Lipid component: 98 % (Chol : 9 %, tri : 82 %, Phosp : < ) % ~ 100 % specifi protein component : · 2% · source : intestine myoglobin · · Heme-containing Function of exogenous triglycerides O2 in binding protein O2 storage , transport o Rapidly released wi muscle damage VLDL (Chol : 22 %, · 52 %, phosp : 18 %) o 1-2 hr post mI Lipid component: 92 % tri : · · Levels normal 10-12 hr post Protein component : 8% · · Detected by immunoassay Source : Liver · Function ELISA , fluroscent , turbidometric , rephelometric of endogenous triglycerides - in transport ↳ sensitive IDL · CTn1 Lipid component : 85 % (Chol : 35 %, tri : 20 %, phosp : 20 % · Comparable (Staus elevated longer · to CK-2 Source : Catabolism of VLDL · · not for early diagnosis Present in low conc. of fasting plasma 04-6 hr post , stays elevated 5-10 days · Precursor of LDL · Fluorescent LDL immunoassay phosp : 23 %) · Lipid component : 79 % (Chol : 47 %, tri : 9 %, · Protein component : 23 % 190-95 % total plasma ApoB-100) · Source : Catabolism of IDL · Functions in cholesterol transport HDL Triglyceride Rich Lipoproteins (Chol: 19 %, tri : 7 %, phosp: 28%) · Chylomicrons: · Lipid component : 50 % 82 % O Lecithin : important functional role in plasma cholesterol ULDL : 52 % esterification catalyzed by LCAT · Protein component : 50 % VLDL · High · source : Liver VLDL directly correlates to high triglyceride level · · fractioned by Further Centrifuge into HDLZ + HDL3 VLDL carries triglycerides in the blood What does creamy layer on serum mean ? multiple Myeloma Caused by · (80 %) high levels of chylomicions Presence of rrine (50 % ) in serum + paraproteins · Bone pain · Anemia · Fatigue · Lipemic Specimen Lutic bone lesions High in triglyceridesO Turbid/milky throughout · · Hypercalcemia IDL · · High in VLDL + Renal insufficiency · 10 % cells in BM plasma ReverseCholesterol transport - nascent HDL Mature HDL - Liver Coronary Heart Disease LDL Risk macrophage · Protects against atherosclerosis High Sensitivity CRP mg/L - · Excess free cholesterd is removed from cells in peripheral tissues 7 10 not consistent w/CHD risk · FC is for excretion ↳ 3 risk returned to the liver in the bile high · FC is generated by the hydrolysis of intracellular Cholesterulester (CE) stores 1-3 normal · < / key molecules: low risk - ATP-binding Cassette protein Al (ABCA1) - - Lecithin : Cholesterol acyltransferase (LCAT) Friedwald Formula to Calculate LDL Conc. in plasma Scavenger (SR-BI) LDL(mg/dL) Total Cholesterol HDL-Triglyceride/S - receptor class B, Type I = - not valid if triglycerides >400mg/dL Uptake of HDL-cholesterol 1) Cholesterol esters taken up from HDL from hepatic HDL receptors ; Beta quantification HDL particles returned to circulation · When friedwald is in appropriate · On EDTA 2) cholesterd esters transferred from HDL to B-100- containing lipoprotein; plasma · Based taken up by liver by receptors for lipoprotein on ultra-polyanion centrifugation · 3) HDL ApoE a 1 006 g/mL = can be recognized by hepatic remnant receptors. 4) Cholesterol returned to liver is either reused or excreted [VLD2) = Stotal) - [d(1. 006gImL] [LDL) = [ d2 1. 006gIm2]-[HDL] HDL-C + LDL - C and CHD C- Reactive Protein + Cardiac Risk Stratification · LDL < 30 mg/dL : Optimal Homocysteine 130-160 mg/dl : Borderline high risk people wl inborn errors of metabolism develop heart disease in teens - ↳ 160 mgld : High risk 0 Levels : > 100Nmol/L < 40 < 5 mol/L -normal HDL mg/dl : High risk : > 40 : Desirable mild elevations associated w/ CHD mg/dL also > 60mgId) : Protective - cause injury to vascular endothelium omediated by inhibition of ROS Type 3 Hyperlipoproteinemia -marker of atherosclerosis · Dysbetalipoproteinemia oprimary genetic defect in removal of intestinal Chylomicrons + hepatic VLDL : Homozygous for ApoEz mutant of ApoE that cannot bind hepatic receptors - Accumulation remnants and - of lipoprotein Cholesterol 1 density <. 00bg/m · B-VLDL or floating B-lipoprotein · Late onset yellow deposit in palm creases - · Lab results ↑ plasma cholesterol + triglycerides - o B-VLDL · Related to TG-rich lipoprotein remnants of hepatic origin t intestinal HDL LDL - + low