Week 3 - Absorbance Spectrophotometry

Question 1

Wavelength (λ)

Answer 1

distance between peaks

expressed in nanometers (nm)

Question 2

Frequency (v)

Answer 2

how many cycles pass in a period of time

Question 3

Velocity (c)

Answer 3

speed of light in a vacuum

c = λ x v

Question 4

Energy (E)

Answer 4

proportional to frequency
E = hv
h = Planck’s constant

Question 5

Transmittance (T)

Answer 5

Light pass through a solution
Ratio of light transmitted by a sample (I) to the light striking the sample (Io) Incident
T = I / Io
Colour we percieve

Question 6

% Transmittance (%T)

Answer 6

T x 100

Question 7

Absorbance (A)

Answer 7

Amount of light absorbed or held back by solution

A = -log T (-log I/Io)

Question 8

How to change T –> A

Answer 8

1. Change T to %T
2. A = log (100 / %T)
   A = 2 - log %T

Question 9

Conditions for Beer-Lambert’s Law (3)

Answer 9

1. Constant path length
2. Monochrome light (single wavelength)
3. Specific concentration ranges (linear)

Question 10

Reagent Blank

Answer 10

Reagent + Water

Question 11

Serum Blank

Answer 11

Serum + Water

Question 12

Patient Sample

Answer 12

Serum + Reagent

Question 13

Working Standard

Answer 13

Reagent + Standard

At least 3

Question 14

Purpose of a sample blank

Answer 14

Account for sample interference

turbidity, hemolysis, extra colour