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104 Cards in this Set

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Describe Lineweaver-Burke plot
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")
Describe the types of inhibition in a Lineweaver-Burke plot
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.
Michaelis-Menton equation
-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
What are the units used to express enzyme activity?
IU = amt enzyme catalyzes conversion of 1 micromole of substrate per minute

1 katal = " 1mole of substrate per sec

1IU = 16.7nanokatals
What are the differences between ALT & AST?
ALT: LIVER & kidney; cytoplasmic; >AST in adults

AST: heart, liver, skel m, kidney, brain, lung, pancreas; cyt & mt (80%)
How many isoenzymes of LDH are there?

LD1,2 in heart, RBCs, kidney
LD3 in lung, spleen, lymphs, panc
LD4,5: liver, skel m
(LD6 - dire finding - hepatic vasc insuff)
Compare LDH isoenzyme concentration between serum & CSF.
Serum: 2>1>3>4>5

CSF: 1>2>3>4>5
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
Where is alkaline phosphatase produced?
Bone (OSTEOBLASTS!), bile ducts, intestine, placenta

Marker of bone-FORMING activity
Eating can factitiously elevate alk phos up to 30%

Decreased in malnutrition

How can we tell the source of alk phos?
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
What is the Regan isoenzyme?
Identical to placental alk phos; Found in 5% ppl with carcinoma
Common clinical situation associated with mildly elevated alk phos
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)
GGT source & use
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
Source & use of 5' nucleotidase
Biliary epithelium
Highest in cholestasis
Confirms liver source of elevated alk phos
Less sens than GGT
Source & use of ammonia
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)


*Requires fresh spec, chilled during transport, NO HEMOLYSIS
Describe bilirubin metabolism
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
Unconjugated hyperbilirubinemia DDx
Blood shunting (cirrhosis)
R heart failure
Gilbert syndrome (not taken up into hepatocyte)
Crigler-Najjar (Not conjugated)
Conjugated hyperbilirubinemia DDx
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
Methods of bilirubin measurement
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)
What is delta-bilirubin?
Conjugated bilirubin covalently linked to albumin. From prolonged conj hyperbili. Cannot be excreted. May persist long after correction of cause.
How does PT measure hepatic function?
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
Which immunoglobulins are useful for diagnosing autoimmune hepatitis? PBC?
ai: polyclonal IgG

PBC: polyclonal IgM
Features of physiologic jaundice in the newborn
DOL 2-3; pks at DOL 4-5
Does not rise at a rate greater than 5mg/dL/day
Red flags for neonatal jaundice
-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
Pathologic correlates of severe neonatal jaundice
Yellowing of subthalamic nucleus, hippocampus, thalamus, globus pallidus, putamen, cerebellar nuclei, cranial nerve nuclei
DDx neonatal hyperbilirubinemia
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)
Conj vs Unconj neonatal hyperbili
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
Use of phototherapy for neonatal jaundice
Only works for UNCONJUGATED bilirubin

Use when bili > 10 before 12h, >12 at 18h, >18 at 24h

**Exchange trx when >20mg/dL
How to test acute HCV?
HCV RNA testing

(anti-HCV antibody only 60% sens)
Transaminase pearls
>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)
Bilirubin pearls
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)
How many isoenzymes of amylase?
3 salivary, 3 pancreatic
Separate by electrophoresis, inhibition tests (salivary sens to inh by wheat germ lectin triticum vulgaris), antibodies
Serum amylase in acute pancreatitis
Rises in 2-24h, returns to nl in 2-3d.

High sens but ~70% spec

Spec of urine amylase and FEamylase is higher
How do you calculate fractional excretion of a compound?
Non-pancreatic causes of high serum amylase
(P-itis usu 250-1000; non-panc 200-500)
Ectopic preg
Bowel Ischemia or Obst
Renal insuff

(Also elevated in panc pseudocyst, IPMN)
Lipase or amylase for acute pancreatitis?

Specific for panc
Elevated for up to 14d
Less reliant on renal clearance
Not affected by triglycerides
Limitations of amylase in dx acute pancreatitis
Not specific
Short window
Affected by renal function
Competitive inhibition by hypertriglyceridemia
What are the Ranson Criteria?
Clues to etiology of pancreatitis
ALT>150 = gallstone (50%sens)

lipase:amylase > 5 = alcoholic (50%sens)

Triglyceride level
Viral serology
How do we test pancreatic exocrine function?
Secretin-CCK (endoscopy)

Fecal fat
Fecal chymotrypsin
Fecal elastase1
Lab evaluation of pancreatic cysts
Name the 3 CK isoenzymes & locations
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.
Types of abnormal CK
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
CK isoenzymes & electrophoresis
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.
What are troponins?
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)
What is the most sensitive cardiac marker?

Earliest marker of AMI.

Also the least cardiospecific.
What are netriuretic peptides?
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
Best markers for non-AMI acute coronary syndrome?
BNP: predicts recurrence & higher likelihood of sudden cardiac death

Troponins: may be helpful

CK-MB, myoglobin: no role
What is ischemia-modified albumin (IMA)?
The amino-terminal binding site of albumin is altered in ischemic conditions. Rises in minutes, cleared in hours. Marker transient ischemia.
Define acute myocardial infarction
Rise & fall of CK-MB or troponin with ischemic symptoms, ECG changes, or coronary artery abnormality.
Cardiac tests in AMI
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
Cardiac enzymes in AMI. How do they change in reperfusion?
Methods of total protein quantification
Nitrogen content (not commonly used)
Refractomerty (interferences)
**Colorimetry in alkaline media (Biuret) (recommended)
Coloriimetry in acid media (for globulins)
Dye binding techniques
Describe the technique of protein electrophoresis. What forces act on each protein?
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)
Which force is greater on gamma globulins in electrophoresis: electromotive or 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.
What are the bands in a normal SPEP, from anode to cathode?
Albumin (most - charge)
Beta (2pks)
Gamma (v slow movement or movement toward cathode)
Is immunoelectrophoresis or immunofixation electrophoresis the preferred method for characterizing a monoclonal band?
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.
Immunoelectrophoresis (IEP)
Immunofixation electrophoresis (IFE)
What is immunotyping?
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.
What is the most abundant protein in human plasma? Function? Fetal equivalent? Half-life?
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)
What is bisalbuminemia?
When a variant allotype is present, 2 peaks can be seen in the albumin band. Benign.
What is the fastest migrating protein on SPEP? Function? Clinical utility? half-life?
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.
How can you tell CSF vs serum SPEP?
-Sharp prealbumin spike in CSF (crosses BBB & excreted by choroid plexus)

-Double Beta-band (transferrin crosses BBB & some is modified to tau protein)
Proteins that migrate to the alpha-1 band?
alpha-1-antitrypsin: major. inactivates proteases. ++ acute phase reactant. (DECREASED in PiZZ phenotype AAT deficiency)

alpha-1-acid glycoprotein: minor. ++ acute phase reactant.
Proteins that migrate to the alpha-2-band?
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)
Proteins that migrate to the beta-band?
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)
What migrates to the second Beta band?
Complement (APR)
Some CRP
Some fibrinogen
Proteins in the gamma region?
- CRP: made in liver.

- immunoglobulins
How is CRP used clinically?
<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
What SPEP pattern is this?
Acute inflammation

Decreased albumin
Increased alpha1 (AAT) & alpha2 (haptoglobin, ceruloplasmin); possible increased beta (complement, fibrinogen)
What SPEP pattern is this?
Chronic inflammation

Similar to acute (increased alpha1&2) but also increased polyclonal gamma globulins
What SPEP pattern is this?
alpha-1 antitrypsin deficiency

(Genotype PiZZ individuals have decreased band. This genotype has <15% normal A1AT, leading to panacinar emphysema & cirrhosis)
What SPEP pattern is this?
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)
What SPEP pattern is this?

Hallmark is beta-gamma bridging**, mainly due to increased IgA.
Also: hypoalbuminemia, blunted alpa1&2 bands.
What SPEP pattern is this?
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
What SPEP pattern is this?

10% of pts with myeloma (likely have free lt chains in urine (BenceJones).
DDx: congenital, lymphoma, nephrotic syndrome, steroids
Which is: normal CSF? abnormal CSF? normal serum?
Top: normal serum

Mid: abnnormal CSF (oligoclonal bands = MS)

Bottom: Normal CSF (** Prominent pre-albumin band & double transferrin/beta band **)
How can UPEP determine source of proteinuria?
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
What are cryoglobulins and what types are there?
= 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.
Palpable purpura, arthralgia, hepatosplenomegaly, lymphadenopathy, anemia, sensorineural deficits, glomerulonephritis. Disease?
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)
Too rapid correction of hyponatremia can lead to?
Central pontine myelinolysis

(too slow correction can lead to cerebral edema)
DDx hyponatremia
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
DDx hypokalemia
GI loss (Uk<30/day)

Renal loss (Uk>30): diuretics, abx, RTA, mineralocorticoid excess, Cushings, CAH

Tran-cellular shifts in alkalosis, correction of DKA
DDx hyperkalemia
artifact (in vitro hemolysis)
renal failure
K+ sparing diuretics (spirono)
adrenal insuff
Which forms of renal tubular acidosis are NOT associated with hyperkalemia?
I & II (low K+)
DDx hypercalcemia
- Primary hyperparathyroidism (inc Ca, dec Phos)
- Tertiary hyperPTH
- Malignancy
- Familial Hypocalciuric Hypercalcemia (CASR gene 3q)
- Drug (vitD)
- Hyperthyroidism, Addison
- Sarcoid
- Paget
-Milk alkali syndrome
Which form of hyperparathyroidism is associated with DECREASED calcium?
Secondary (peripheral resistance to PTH)

(Primary: 90% PTH adenoma; MEN1, 2A)
(Tertiary: post-renal transplant)
What is humoral hypercalcemia of malignancy?
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.
Symptoms hypercalcemia
kidney stones
peaked T waves on EKG
Types of hyperPTH?
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)
Elevated Ca++ and Phosphate?
Hypervitaminosis D

(sarcoid granulomas can activate Vitamin D!)
Sx hypercalcemia?
Lethargy, nephrolithiasis, hyporeflexia, N, V, constip


Risk of pancreatitis & PUD

Long term with hyperphosphate: CALCIPHYLAXIS (Ca of vessel walls & soft tissue)
Besides clinical history, how to distinguish cause of hypercalcemia?
HyperPTH vs malignancy (PTHrP)

-second gen tests can detect N-terminal PTH or intact PTH (the only active forms), and can ID PTHrP
Total CA++ vs ionized?
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...
Sx hypocalcemia?
neurologic excitability
muscle spasms
QT prolongation
Tetany, laryngeal spasm
Anion gap calculation
AG = Na - (Cl + HCO3)

Normal is < 12
How does albumin affect the anion gap?
Hypoalbuminemia can MASK an anion gap

Corrected AG = AG + 2.5(4-alb)
Causes of increased anion gap:
Lactic Acidosis
Ethylene glycol
Osmol gap calculation
OG = measured osms - (2Na + glu/18 + BUN/2.8)

Nl < 10
Causes of increased osmol gap:
with metabolic acidosis:
methanol, propylene glycol, diethylene glycol, paraldehyde, ethanol

isopropyl alcohol, glycerol, sorbitol, mannitol, acetone, ethanol (sometimes)
Compensation of HCO3 for each 1 mmHg change in pCO2:
MAcid: 1.3
MAlk: 0.6
Resp: 0.1 acute; 0.4 chronic
FENa calculation
FENa < 1%
PRERENAL disease
Usually hypovolemia
Hepatorenal syndrome
Acute glomerulonephropathy
FENa > 3%
Kidney damage
(ATN or obstruction)