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104 Cards in this Set
- Front
- Back
Describe Lineweaver-Burke plot
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Plot of enzyme kinetics
y-axis: 1/velocity y-interecept: 1/Vmax x-axis: 1/substrate concentration x-intercept: -1/Km (Km is the concentration of substrate that leads to half-maximal velocity; aka "Michaelis-Menton constant") |
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Describe the types of inhibition in a Lineweaver-Burke plot
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Competitive inhibition: Vmax does not change. Competes with substrate for binding but can be overcome as increase substrate concentration. x-intercept changes.
Non-competitive inhibition: Vmax changes. Cannot be overcome by increasing substrate concentration. y-intercept changed; x-intercept does not. Uncompetitive inhibition: Rare. Binds enzyme-substrate complex & stabilizes. changes BOTH x&y intercepts. |
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Michaelis-Menton equation
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-Measurement of enzyme activity
-Rate of enzyme activity varies with substrate concentration, until saturated (Vmax) -At saturation, Vmax varies only with enzyme concentration -Km = substrate concentration at half-Vmax |
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What are the units used to express enzyme activity?
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IU = amt enzyme catalyzes conversion of 1 micromole of substrate per minute
1 katal = " 1mole of substrate per sec 1IU = 16.7nanokatals |
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What are the differences between ALT & AST?
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ALT: LIVER & kidney; cytoplasmic; >AST in adults
AST: heart, liver, skel m, kidney, brain, lung, pancreas; cyt & mt (80%) |
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How many isoenzymes of LDH are there?
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5
LD1,2 in heart, RBCs, kidney LD3 in lung, spleen, lymphs, panc LD4,5: liver, skel m (LD6 - dire finding - hepatic vasc insuff) |
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Compare LDH isoenzyme concentration between serum & CSF.
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Serum: 2>1>3>4>5
CSF: 1>2>3>4>5 |
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What are the possible causes of serum LD1>LD2?
Elevation of LD4&5? Elevation of LD1&5? |
MI, hemolysis, renal infarction
Liver damage, skel muscle Acute MI + liver congestion OR chronic ETOH + liver damage + megaloblastic anemia |
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Where is alkaline phosphatase produced?
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Bone (OSTEOBLASTS!), bile ducts, intestine, placenta
Marker of bone-FORMING activity Eating can factitiously elevate alk phos up to 30% Decreased in malnutrition m>f |
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How can we tell the source of alk phos?
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Has 4 isoenzymes by e-phoresis that have unique degrees of inactivation by heat, urea, & l-phenylalanine.
** BONE BURNS: 90% inactivation of bone alk phos with heating; 0% of placental; 50% biliary |
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What is the Regan isoenzyme?
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Identical to placental alk phos; Found in 5% ppl with carcinoma
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Common clinical situation associated with mildly elevated alk phos
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Pregnancy
CHF Hyperthyroidism Drug (ibuprofen, acetaminophen) anti-mitochondrial antibodies (middle aged women) ** Sensitive marker for hepatic mets Suspect bone origin without concomitant elevation GGT or 5'nucleotidase (PAGETS, hyperPTH, osteomalacia) |
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GGT source & use
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Biliary epithelial cell (v sensitive for biliary injury); also hepatocytes
Used to confirm elevated alk phos is coming from biliary tree Also elevated with toxins (EtOH, warfarin) Can be used as marker of alcohol consumption M >> F Af Am >> W |
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Source & use of 5' nucleotidase
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Biliary epithelium
Highest in cholestasis Confirms liver source of elevated alk phos Less sens than GGT |
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Source & use of ammonia
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Skel muscle & gut
Made by bacteria in gut from protein; excreted as urea by liver in urine Hyperammonia = liver failure (inless inborn error urea metab) Neurotoxic *Requires fresh spec, chilled during transport, NO HEMOLYSIS |
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Describe bilirubin metabolism
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Unconj = Indirect = water insol
Binds albumin --> liver --> glucuronidation to Conj (Direct) --> excretion into bile --> converted to urobilinogen by intest bacteria (some to kidney) --> colonic bact converts some to brown pigment ** BILIRUBINURIA = CONJ HYPERBILI as unconj never in urine |
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Unconjugated hyperbilirubinemia DDx
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Hemolysis
Blood shunting (cirrhosis) R heart failure Gilbert syndrome (not taken up into hepatocyte) Drug(rifampin) Crigler-Najjar (Not conjugated) Hypothyroid |
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Conjugated hyperbilirubinemia DDx
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Mechanical obstruction (PBC, PSC, stricture, stone) (CHOLESTATIC jaundice)
**>3x upper nl alk phos, elevated cholesterol, pruritis, mild elev transaminases HEPATOCELLULAR jaundice (Not secreted: Dubin-Johnson, Hepatitis, Endotoxin/sepsis, Pregnany/Estrogen, drug-E, cyclosporine) **>3x upper nl transaminases, nl cholesterol, no pruritis, mild elev alk phos |
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Methods of bilirubin measurement
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1)Diazo-colorimetric methods: Measures conjugated; with accelerator can measure total (unconj is then estimated)
2) Direct spectrophotometry: Measures total bili; absorbance at 455 nm - that at 575 nm (hgb interference) |
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What is delta-bilirubin?
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Conjugated bilirubin covalently linked to albumin. From prolonged conj hyperbili. Cannot be excreted. May persist long after correction of cause.
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How does PT measure hepatic function?
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Most coagulation factors synthesized in hepatocytes (shortest T 1/2 FVII 12 hours)
Impaired bile secretion can lead to vit K deficiency BEST indicator of px in acute hepatic injury |
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Which immunoglobulins are useful for diagnosing autoimmune hepatitis? PBC?
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ai: polyclonal IgG
PBC: polyclonal IgM |
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Features of physiologic jaundice in the newborn
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DOL 2-3; pks at DOL 4-5
Does not rise at a rate greater than 5mg/dL/day |
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Red flags for neonatal jaundice
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-Jaundice within first 24h
-Rising bilirubin beyond 1wk -Persistent jaundice >10d -Total bili > 12mg/dL -Single-day increase >5mg/dL -Conjugated bili > 2mg/dL **severe: HDN, sepsis. Causes kernicterus |
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Pathologic correlates of severe neonatal jaundice
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Yellowing of subthalamic nucleus, hippocampus, thalamus, globus pallidus, putamen, cerebellar nuclei, cranial nerve nuclei
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DDx neonatal hyperbilirubinemia
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1st 24h: erythroblastosis fetalis, hemorrhage, sepsis, TORCH infxn
3-7d: sepsis (UTI) >1wk: br milk jaundice, sepsis, biliary atresia, CF, Alagille synd (congen paucity bds), neonatal hepatitis, galactosemia, inherited hemol anemia (PK def, HS, G6PD def) |
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Conj vs Unconj neonatal hyperbili
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Unconj: physiologic, br milk, polycythemia, hemolysis, bowel obst, inherited (GILBERT, C-N)
Conj: Biliary obst, sepsis, TORCH, hepatis, metabolic disorders, inherited (D-J, ROTOR), TPN |
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Use of phototherapy for neonatal jaundice
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Only works for UNCONJUGATED bilirubin
Use when bili > 10 before 12h, >12 at 18h, >18 at 24h **Exchange trx when >20mg/dL |
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How to test acute HCV?
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HCV RNA testing
(anti-HCV antibody only 60% sens) |
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Transaminase pearls
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>1000x ULN = acute hepatic inj from ischemia or toxin (not viral)
AST > 3000 U/L = TOXIN AST:ALT ratio >2 in toxic, ischemic, EtOH; <1 in viral Poorly reflect degree of hepatic injury (vs PT, bili) |
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Bilirubin pearls
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jaundice more common in HAV & alcoholic hepatitis > HBV, HCV; rare in kids with viral hepatitis
Bilirubin peaks after transaminases fall in all forms of acute hepatic injury Bili > 15mg/dL indicative severe liver inj & poor px Acute hepatic injury mimics obstruction (direct > indirect) |
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How many isoenzymes of amylase?
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6
3 salivary, 3 pancreatic Separate by electrophoresis, inhibition tests (salivary sens to inh by wheat germ lectin triticum vulgaris), antibodies |
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Serum amylase in acute pancreatitis
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Rises in 2-24h, returns to nl in 2-3d.
High sens but ~70% spec Spec of urine amylase and FEamylase is higher |
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How do you calculate fractional excretion of a compound?
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Non-pancreatic causes of high serum amylase
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(P-itis usu 250-1000; non-panc 200-500)
DKA PUD Cholecystitis Ectopic preg Salpingitis Bowel Ischemia or Obst Renal insuff Opioids (Also elevated in panc pseudocyst, IPMN) |
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Lipase or amylase for acute pancreatitis?
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LIPASE
Specific for panc Elevated for up to 14d Less reliant on renal clearance Not affected by triglycerides |
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Limitations of amylase in dx acute pancreatitis
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Not specific
Short window Affected by renal function Competitive inhibition by hypertriglyceridemia |
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What are the Ranson Criteria?
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Clues to etiology of pancreatitis
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ALT>150 = gallstone (50%sens)
lipase:amylase > 5 = alcoholic (50%sens) Triglyceride level Ca++ Viral serology |
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How do we test pancreatic exocrine function?
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Secretin-CCK (endoscopy)
Fecal fat Fecal chymotrypsin Fecal elastase1 |
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Lab evaluation of pancreatic cysts
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Name the 3 CK isoenzymes & locations
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CK-BB: BRAIN, bladder, stomach, prostate... all tissues. Fastest on e-phoresis
CK-MM: Skel m, cardiac m. Slowest. (Serum CK is usu 100% CK-MM, from skel m) CK-MB: cardiac m (30%), skel m (1%). In nl pts, the circulating CK-MB is from skel m. |
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Types of abnormal CK
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Macro-CK: CK-Ig complex. Healthy elderly f. Migrates between MM-MB
Mitochondrial CK: Poor px; seen in pts with advanced malignancies. Migrates with MM |
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CK isoenzymes & electrophoresis
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CK-BB: BRAIN, bladder, stomach, prostate... all tissues. Fastest on e-phoresis
CK-MM: Skel m, cardiac m. Slowest. (Serum CK is usu 100% CK-MM, from skel m) CK-MB: cardiac m (30%), skel m (1%). In nl pts, the circulating CK-MB is from skel m. |
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What are troponins?
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Group pf enzymes (T,I,C) that mediate actin-myosin interactions of muscle contraction.
Cardiac troponin I tested in lab. Cardiac specific (not altered in skel m injury - unlike CK-MB, myoglobin) Immediate release with infarction (4-8h) as well as sustained release (10-14d) |
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What is the most sensitive cardiac marker?
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Myoglobin.
Earliest marker of AMI. Also the least cardiospecific. |
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What are netriuretic peptides?
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Cause vasodilation & sodium excretion.
3 types: ANP, BNP (ventricular myocytes; correlates with v wall tension; rapid degradation), CNP BNP correlates with fluid status; EXCELLENT MARKER FOR CHF |
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Best markers for non-AMI acute coronary syndrome?
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BNP: predicts recurrence & higher likelihood of sudden cardiac death
Troponins: may be helpful CK-MB, myoglobin: no role |
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What is ischemia-modified albumin (IMA)?
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The amino-terminal binding site of albumin is altered in ischemic conditions. Rises in minutes, cleared in hours. Marker transient ischemia.
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Define acute myocardial infarction
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Rise & fall of CK-MB or troponin with ischemic symptoms, ECG changes, or coronary artery abnormality.
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Cardiac tests in AMI
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Troponin: Single+ highly susp; double+ better; (-) result in days excludes. If+, still need CK-MB to determine acute/resolving.
CK-MB: rise in 3-6h, pk 24h, nl in 72h. Great sens but less spec. Myoglobin: rapid release, not spec. Cardiac reperfusion: all markers peak EARLIER |
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Cardiac enzymes in AMI. How do they change in reperfusion?
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Methods of total protein quantification
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Nitrogen content (not commonly used)
Refractomerty (interferences) **Colorimetry in alkaline media (Biuret) (recommended) Coloriimetry in acid media (for globulins) Dye binding techniques Spectrophotometry |
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Describe the technique of protein electrophoresis. What forces act on each protein?
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Movement of proteins due to electrical potential
Charge applied across solid support & fluid buffer. SS (-) charge so is drawn to + POLE (ANODE); fluid buffer drawn to - POLE (CATHODE) 2 forces act on each protein: -ELECTROMOTIVE: Pulled toward opposite charge (usu to ANODE) -ENDOSMOTIC: flow with the fluid buffer (toward CATHODE) |
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Which force is greater on gamma globulins in electrophoresis: electromotive or endosmotic?
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Endosmotic.
Gamma globulins have a weak negative charge; the endosmotic force (toward CATHODE) outweighs the electromotive force. Not true with most other proteins, which are more negatively charged and this driven to the + ANODE. |
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What are the bands in a normal SPEP, from anode to cathode?
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Albumin (most - charge)
Alpha1 Alpha2 Beta (2pks) Gamma (v slow movement or movement toward cathode) |
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Is immunoelectrophoresis or immunofixation electrophoresis the preferred method for characterizing a monoclonal band?
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Immunofixation electrophoresis (IFE): simpler to interpret
IEP: run pt & nl serum in adjacent troughs, then run antisera, precipitation arcs form. should be symmetric. IFE: pt serum in wells on gel, e-phoresis, antisera applied, stained. |
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Immunoelectrophoresis (IEP)
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Immunofixation electrophoresis (IFE)
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What is immunotyping?
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aka immunosubtraction
used with capillary electrophoresis to identify M protein. serum incubated with antibodies; supernatants removed & e-phoresed - can see which reagents removed abnormal spike. |
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What is the most abundant protein in human plasma? Function? Fetal equivalent? Half-life?
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Albumin (2/3 total plasma protein)
Carries substances; Assessment of nutritional status. alpha-fetoprotein is fetal equivalent (but migrates to alpha-1) t1/2 = 17days also: serum decrements in protein-losing conditions; glycosylated >8% in diabetes; negative acute phase reactant (decreases in inflammatory conditions) |
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What is bisalbuminemia?
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When a variant allotype is present, 2 peaks can be seen in the albumin band. Benign.
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What is the fastest migrating protein on SPEP? Function? Clinical utility? half-life?
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Prealbumin. Sparse though and not traditionally seen without high resolution.
Binds T3 (transthyretin), T4 (thyroxine-binding prealbumin), retinol-binding protein:vitaminA complex. Assessment of nutritional status t1/2 = 48h also: rare variant with very high affinity for thyroxine (elevated total T4 in euthyroid pt); heparinzed pts may have false "prealbumin band" with altered beta-lipoprotein; elevated prealb in chr EtOH, corticosteroid use; negative acute phase reactant. |
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How can you tell CSF vs serum SPEP?
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-Sharp prealbumin spike in CSF (crosses BBB & excreted by choroid plexus)
-Double Beta-band (transferrin crosses BBB & some is modified to tau protein) |
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Proteins that migrate to the alpha-1 band?
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alpha-1-antitrypsin: major. inactivates proteases. ++ acute phase reactant. (DECREASED in PiZZ phenotype AAT deficiency)
alpha-1-acid glycoprotein: minor. ++ acute phase reactant. |
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Proteins that migrate to the alpha-2-band?
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alpha-2-macroglobulin: elevated in liver & renal dz (lrg size not excreted well)
Ceruloplasmin: copper trx. decreased in Wilson dz. Also in hepatic failure, malnutrition, Menke synd (AR, "kinky hair synd"). Acute phase reactant (APR) Haptoglobin: Binds free hgb (DECREASED in hemolysis = sensitive marker). APR. 2 alleles = 3 haplotypes (2-2 phenotype risk factor for CV disease in diabetics) |
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Proteins that migrate to the beta-band?
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TRANSFERRIN: Transports Fe+++ (30% sat'd); INCREASED in iron def (mimics M-protein), pregnancy, estrogen thx. NEGATIVE APR. Crosses BBB (modifies some to TAU protein - double pk in CSF)
Fibrinogen (usually not in serum unless incomplete clot; straddles beta-gamma interface; present in dysfibrinogenemia, APL synd, liver dz, vit K def, heparin use) |
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What migrates to the second Beta band?
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IgA
Complement (APR) Some CRP Some fibrinogen |
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Proteins in the gamma region?
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- CRP: made in liver.
- immunoglobulins |
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How is CRP used clinically?
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<3mg/L : normal
3-10: cellular stress >10: active inflammation low-level elevation predicts poor outcome after CV events as well as in healthy individuals |
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What SPEP pattern is this?
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Acute inflammation
Decreased albumin Increased alpha1 (AAT) & alpha2 (haptoglobin, ceruloplasmin); possible increased beta (complement, fibrinogen) |
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What SPEP pattern is this?
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Chronic inflammation
Similar to acute (increased alpha1&2) but also increased polyclonal gamma globulins |
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What SPEP pattern is this?
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alpha-1 antitrypsin deficiency
(Genotype PiZZ individuals have decreased band. This genotype has <15% normal A1AT, leading to panacinar emphysema & cirrhosis) |
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What SPEP pattern is this?
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Nephrotic syndrome
Massive loss of small proteins (only albumin in minimal change dz). Leads to "dimming" of all the bands, esp albumin, with SPARING of alpha2 (large protein alpha-2 macroglobin) |
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What SPEP pattern is this?
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Cirrhosis
Hallmark is beta-gamma bridging**, mainly due to increased IgA. Also: hypoalbuminemia, blunted alpa1&2 bands. |
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What SPEP pattern is this?
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Monoclonal gammopathy
aka paraproteinemia Prominent M-SPIKE, usu in gamma but can be in beta or alpha2. Requires IFE to characterize/ DDx: plasma cell dyscrasia, Waldenstrom/LPL, CLL, AL amyloidosis Look for cryoglobulins in pts with M-proteins |
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What SPEP pattern is this?
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Hypogammaglobulinemia
10% of pts with myeloma (likely have free lt chains in urine (BenceJones). DDx: congenital, lymphoma, nephrotic syndrome, steroids |
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Which is: normal CSF? abnormal CSF? normal serum?
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Top: normal serum
Mid: abnnormal CSF (oligoclonal bands = MS) Bottom: Normal CSF (** Prominent pre-albumin band & double transferrin/beta band **) |
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How can UPEP determine source of proteinuria?
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Glomerular pattern: Very large & very small proteins retained. Thus, strong albumin, alpha1(AAT) & beta (transferrin)
Tubular pattern: Impaired tubular reabsorption of LMW proteins such as alpha1macroglobin, beta2microglobin, and light chains. Thus, weak albumin band and strong alpha1 & beta bands. Overflow proteinuria: Usu monoclonal light chain (BenceJones); also myoglobin or hgb |
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What are cryoglobulins and what types are there?
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= immunoglobulins that reversibly precipitate at low temperatures
I: monoclonal Ig assoc with MM or Waldenstrom II: mixture monoclonal IgM & polyclonal IgG. The IgM has RF activity (anti-IgG). **Most common III: Mixture of 2 polyclonal Ig. |
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Palpable purpura, arthralgia, hepatosplenomegaly, lymphadenopathy, anemia, sensorineural deficits, glomerulonephritis. Disease?
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Mixed cryoglobulinemia (Types II&III)
DDx: HCV**, chronic infection, lymphoproliferative, SLE, liver dz. Usu females 30-50. Vasculitis; MPGN. EM: large subendothelial immune complex deposits. Thx: steroids, plasmapheresis, interferon (for HCV) |
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Too rapid correction of hyponatremia can lead to?
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Central pontine myelinolysis
(too slow correction can lead to cerebral edema) |
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DDx hyponatremia
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SIADH (urine Na >20, euvolemia, nl renal fxn, no recent diuretics)
Pseudohyponatremia (serum osm >280: hypergly, hyperprot, hyperlipid) + Hypovolemia: renal: UNa>30; extrarenal: UNa<30 +Euvolemia: SIADH, psych, ADH-like drugs +Hypervolemia: CHF, cirrhosis, nephrotic syndrome |
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DDx hypokalemia
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GI loss (Uk<30/day)
Renal loss (Uk>30): diuretics, abx, RTA, mineralocorticoid excess, Cushings, CAH Tran-cellular shifts in alkalosis, correction of DKA |
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DDx hyperkalemia
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artifact (in vitro hemolysis)
acidosis renal failure K+ sparing diuretics (spirono) adrenal insuff rhabdo |
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Which forms of renal tubular acidosis are NOT associated with hyperkalemia?
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I & II (low K+)
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DDx hypercalcemia
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- Primary hyperparathyroidism (inc Ca, dec Phos)
- Tertiary hyperPTH - Malignancy - Familial Hypocalciuric Hypercalcemia (CASR gene 3q) - Drug (vitD) - Hyperthyroidism, Addison - Sarcoid - Paget -Milk alkali syndrome |
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Which form of hyperparathyroidism is associated with DECREASED calcium?
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Secondary (peripheral resistance to PTH)
(Primary: 90% PTH adenoma; MEN1, 2A) (Tertiary: post-renal transplant) |
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What is humoral hypercalcemia of malignancy?
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Malignancy produces hypercalcemia WITHOUT bone mets
RCC, HCC, SCC, PG, islet cell tumors PTHrp related. HIGHLY suggested when increased nephrogenous cAMP in presence of normal PTH. |
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Symptoms hypercalcemia
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kidney stones
lethargy hyporeflexia n/v/constipation peaked T waves on EKG |
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Types of hyperPTH?
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Primary: PTH adenoma (90%), 4-gland hyperplasia (9%), ca
Secondary: Peripheral resistance to PTH; LOW CA++ (usu chronic renal failure. also Vit D def). (BROWN TUMORS OF BONE - persistent hyperPTH activates osteoclasts) Tertiary: HyperPTH that persists post-renal transplant (autonomous PTHs) |
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Elevated Ca++ and Phosphate?
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Hypervitaminosis D
(sarcoid granulomas can activate Vitamin D!) |
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Sx hypercalcemia?
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Lethargy, nephrolithiasis, hyporeflexia, N, V, constip
PEAKED T WAVES EKD Risk of pancreatitis & PUD Long term with hyperphosphate: CALCIPHYLAXIS (Ca of vessel walls & soft tissue) |
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Besides clinical history, how to distinguish cause of hypercalcemia?
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HyperPTH vs malignancy (PTHrP)
-second gen tests can detect N-terminal PTH or intact PTH (the only active forms), and can ID PTHrP |
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Total CA++ vs ionized?
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50% CA++ is bound to albumin
Ionized/free is biologically active Acidosis increases free CA++ Measuring ionized CA++ is tricky. Arterial, no air exposure, kept cool delivered rapidly... |
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Sx hypocalcemia?
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neurologic excitability
muscle spasms QT prolongation Tetany, laryngeal spasm |
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Anion gap calculation
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AG = Na - (Cl + HCO3)
Normal is < 12 |
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How does albumin affect the anion gap?
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Hypoalbuminemia can MASK an anion gap
Corrected AG = AG + 2.5(4-alb) |
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Causes of increased anion gap:
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Methanol
Uremia Ketoacidosis paraldehyde Lactic Acidosis Ethylene glycol Salicylate |
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Osmol gap calculation
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OG = measured osms - (2Na + glu/18 + BUN/2.8)
Nl < 10 |
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Causes of increased osmol gap:
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with metabolic acidosis:
methanol, propylene glycol, diethylene glycol, paraldehyde, ethanol without: isopropyl alcohol, glycerol, sorbitol, mannitol, acetone, ethanol (sometimes) |
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Compensation of HCO3 for each 1 mmHg change in pCO2:
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MAcid: 1.3
MAlk: 0.6 Resp: 0.1 acute; 0.4 chronic |
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FENa calculation
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FENa < 1%
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PRERENAL disease
Usually hypovolemia Hepatorenal syndrome Acute glomerulonephropathy CHF |
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FENa > 3%
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Kidney damage
(ATN or obstruction) |