PRAC 1 DATA ANALYSIS

STEP 1 INPUT ABSORBANCE RAW DATA OBTAINED FROM THE SPECTROPHOTOMETER

RAW MICROPLATE DATA

1 2 3 4 5 6
A 0.347 0.362 0.34 0.352 0.355 0.352
B 0.34 0.359 0.522 0.492 0.658 0.626
C 0.702 0.854 0.514 0.529 1.583 1.593

STEP 2 CALCULATE AVERAGE ABSORBANCE OF BLANK SAMPLES (ROW A)

1 2 3 4 5 6
A 0.347 0.362 0.34 0.352 0.355 0.352

STEP 3 CALCULATE CORRECTED ABSORBANCE DATA BY SUBTRACTING THE AVERAGE OF B

1 2 3 4 5 6
B -0.0141667 0.00483333 0.167833333 0.137833 0.303833 0.271833
C 0.34783333 0.49983333 0.159833333 0.174833 1.228833 1.238833

STEP 4 CALCULATE AVERAGES FOR YOUR CORRECTED REPLICATES (ROWS B & C in STEP 3

Well Number AVERAGE Tip: use Excel Average formula

B1 and B2 (No Protein) -0.005
B3 and B4 (5µL of Protein B) 0.1528
B5 and B6 (10µL of Protein B) 0.2878
B7 and B8 (5µL of Protein A) 0.6328
B9 and B10 (10µL of Protein A) 0.9118
B11 and B12 (20µL of Protein A) 1.1778
C1 and C2 (Unknown A) 0.4238
C3 and C4 (Unknown B) 0.1673
C5, C6, C7 and C8 (Unknown C) 1.1541

STEP 5 PREPARE DATA FOR PLOTTING A STANDARD CURVE

Protein AVERAGE
µg Reading Tip: those values will be automatically insterted from STEP 4
0 -0.00466667
1 0.152833333
 2 0.287833333
5 0.632833333
10 0.911833333
20 1.177833333

STEP 6 PLOT THE ABOVE VALUES AND ESTIMATE A LINE OF BEST FIT TO OBTAIN THE STAN

AVERAGE
1.4

1.2
f(x) = - 0.0035796102x^2 + 0.1287250053x + 0.0273572038
R² = 0.994324389

0.8

0.6

0.4

0.2

0
0 5 10 15

-0.2

STEP 7 USING THE STANDARD CURVE SOLVE THE EQUATION FROM THE LINE OF BEST FIT

Unknown AVERAGE Protein

Sample Reading µg
Sample A 0.423833333 3.4045 Tip: AVERAGE values will be automatically insterted from STEP 4
Sample B 0.167333333 1.12252
Sample C 1.154083333 15.31608
Linear line of best fit has equation as follows: y = ax+c where solving for x determines the protein amou
Polynominal line of best fithas equation as follows: y = ax2 + bx + c, where solving for x determines the
ECTROPHOTOMETER

OPLATE DATA
7 8 9 10 11 12
0.35 0.356 0.362 0.357 0.365 0.352 Tip: require your attention
0.968 1.006 1.282 1.25 1.473 1.591
1.43 1.427

7 8 9 10 11 12 AVERAGE Tip: use Excel Average formula

0.35 0.356 0.362 0.357 0.365 0.352 0.354167

TING THE AVERAGE OF BLANK SAMPLES (STEP 2) FROM YOUR ACTUAL RESULTS (ROWS B & C)

7 8 9 10 11 12
0.613833 0.651833 0.927833 0.895833 1.118833 1.236833 Tip: those values will be automatically calculated from STEP
1.075833 1.072833

S (ROWS B & C in STEP 3)

xcel Average formula

rom STEP 4
FIT TO OBTAIN THE STANDARD CURVE. FIT THE GRAPH IN THE BELOW BOX

GE

A -0.0036
B 0.1287
C

15 20 25

M THE LINE OF BEST FIT TO ESTIMATE TOTAL PROTEIN CONCETRATION (EXPRESSED AS TOTAL MICROGRAMS

C
atically insterted from STEP 4 0.0274 -0.396433
0.0274 -0.139933
0.0274 -1.126683
 determines the protein amount of the sample
e solving for x determines the protein amount of the sample
our attention

WS B & C)

matically calculated from STEP 1 & 2

AS TOTAL MICROGRAMS OF PROTEIN)