Unit 3 - Redone Flashcards
Primary Structure
Sequence of amino acids
Secondary Structure
Amino acid chains with a more complex structure
Tertiary Structure
Complex folding of polypeptide chains
Quaternary Structure
2+ polypeptide chains
Simple vs Conjugated Proteins
Conjugated proteins have a non-amino chemical associated with them, Simple proteins don’t
215nm
Peptide bonds
280nm
Aromatic rings in amino acids
Tyrosine
Tryptophan
Phenylalanine
Biuret reagent
Copper sulfate + NaOH
Do amino acids and dipeptides react with biuret? Why or why not?
No, require at least TWO peptide BONDS.
What interferants can interfere with biuret measurement?
Hemolysis and icterus
Biuret color change in the presence of protein
Blue to Violet
Total serum protein amount
6-8 g/dL
Biuret cannot be used with what specimen?
Urine, CSF
Biuret is insensitive for what kinds of proteins?
Globulins and other proteins that aren’t albumin
When would biuret’s insensitivity to other proteins be an issue?
When multiple myeloma present
Albumin measurement
Bromcresol green
Bromcresol purple
What % of plasma protein is albumin?
50%
Globulins definition
All plasma proteins NOT albumin
TP formula
Albumin + Globulins
Globulins calculation
Total protein - Albumin = Globulins
A/G Ratio
Albumin / Globulins
Albumins reference intervals
3.5-5 g/dL
Globulins reference intervals
2.5-3 g/dL
Urine protein reference intervals
1-14 mg/dL
or
< 100 mg/24hr
CSF Protein reference intervals
15-45 mg/dL
Buffer of Electrophoresis pH; why is it that?
8.6, Alb & Globs net negative charge so they will move to their isoelectric points
Which protein has the biggest net negative charge and thus moves fastest?
Albumins
Formula for urine total protein measurement per 24 hours
look it up
Anode
Positive pole; Anions move towards it
Cathode
Negative pole; Cations move towards it
Heavy application point can be caused by
Thawed specimen with precipitated protein on it
Which end of the medium is the application point?
Towards the cathode end (negative end)
Lowest net negative charge
Globulins
Highest net negative charge
Albumin
Most anodal band
Albumin, the fastest
Most cathodal band
Globulins, the slowest
Frictional Coefficient: Larger pores
Separation based on size AND charge
Larger and more complex molecules move slower
Frictional Coefficient: Smaller pores
Separation based on charge only
What Is the electrolyte present in the medium that is required for electrons to flow through the medium?
Sodium barbital
Support Media and their uses
Paper - Historic & Awful
Cellulose Acetate - Historic
Agarose - Common, separation based on charge
Polyacrylamide - Specialized, separation based on charge and size
What causes smile gel
Temperature
Wick flow
Evaporation, proteins make a trail
How is wick flow fixed
Keep a lid on
Cool the system
Change buffer often/use fresh buffer
Electroendosmosis
Cations bump into anions like gamma globulins as they move towards the cathode and cause the gamma globulins to move backwards
Why does the albumin band look fat
Tracer dyes bind to a portion of the albumin
Two albumin bands can be due to
Drugs that bind to albumin
or
Genetic bisalbuminemia
Complement band on SPEP
In fresh specimen
Faint line in second Beta region
Fibrinogen band
Plasma or specimen that didn’t clot
Between beta and gamma (after second band)
How to fix fibrinogen band
Add thrombin so that specimen clots
What does hemolysis do to SPEP
Joins alpha-2 and beta bands
Where is CRP seen on SPEP?
Gamma region near application point
Serum proteins are produced where?
Mostly in the liver, but not the antibodies
