Week 5: Electrophoresis

zone EP

protein fractions separate into distinct bands

densitometry

1. EP recording method

2. zone identification and quantification

electrophoretogram (SEM serum protein EP)

1. result from densitometry

2. pattern of separated protein peaks

protein electrophoretic mobility factors

1. protein molecule

2. temperature

3. electric field strength

4. support medium type

an acidic solution makes proteins __________, while a basic solution makes proteins ___________

positive, negative

buffer used for serum protein and hemoglobin EP

1. barbital or tris buffer

2. pH 8.6-8.8

buffer and gel used for abnormal hemoglobin EP

1. citrate agarose gel

2. citrate buffer

3. pH 5-6

types of EP support media

1. cellulose acetate

2. agarose gel

3. polyacrylamide gel

electroendosmosis

positively charged support medium and buffer flow cause weakly charged proteins to migrate to the negative pole

electroendosmosis results in EP __________

artifacts

protein stains

1. ponceau S

2. coomassie brilliant blue

3. amido black

lipoprotein stains

1. oil red O, fat red

2. sudan black

a densitometer is a modified ______________

spectrophotometer

a densitometer directs _______________ light onto stained bands

monochromatic

in densitometry, absorbance is proportional to _____________

concentration

applications of electrophoresis

1. SEM

2. DNA isolation

3. forensics

4. capillary EP