BIT 410/510 Lab 9
Affinity Chromatography, in vitro Manipulation & Bradford
Assay
, Lab 9 Order of Operations
❖ Reminders/Recap
❖ Protein Purification
❖ Affinity chromatography
❖ in vitro Protein Manipulation
❖ Testing calcium responsiveness of GST:CaMPARI
❖ Quantification
❖ Bradford assay
, Reminders!
ES2 due in two weeks (week of 11/6) - 12:50 before lab
●Lab activities: SDS-PAGE/Western Blot, affinity chromatography, Bradford analy
Ca2+ responsiveness of purified CaMPARI
●Figure 1 (A-C): Image of SDS-PAGE (A), Membrane after transfer (B), and West
blot chemiluminescence (C) of the induction test samples
●Table 1: Values from your ImageJ analysis (Average Normalized Protein Value a
Fold Change)
●Figure 2 (A-C): UV light images of (A) affinity chromatography fractions, (B) p
CaMPARI fraction after addition of calcium, and (C) addition of EGTA
●Figure 3: Graph of Bradford standard curve
●Table 2: Total protein of the affinity chromatography fractions (Bradford results)
Executive Summary Guidelines
Affinity Chromatography, in vitro Manipulation & Bradford
Assay
, Lab 9 Order of Operations
❖ Reminders/Recap
❖ Protein Purification
❖ Affinity chromatography
❖ in vitro Protein Manipulation
❖ Testing calcium responsiveness of GST:CaMPARI
❖ Quantification
❖ Bradford assay
, Reminders!
ES2 due in two weeks (week of 11/6) - 12:50 before lab
●Lab activities: SDS-PAGE/Western Blot, affinity chromatography, Bradford analy
Ca2+ responsiveness of purified CaMPARI
●Figure 1 (A-C): Image of SDS-PAGE (A), Membrane after transfer (B), and West
blot chemiluminescence (C) of the induction test samples
●Table 1: Values from your ImageJ analysis (Average Normalized Protein Value a
Fold Change)
●Figure 2 (A-C): UV light images of (A) affinity chromatography fractions, (B) p
CaMPARI fraction after addition of calcium, and (C) addition of EGTA
●Figure 3: Graph of Bradford standard curve
●Table 2: Total protein of the affinity chromatography fractions (Bradford results)
Executive Summary Guidelines
