Limit of Detection and Lower Limit of the Measuring Interval

References: CLSI C62-A (October 2014), CLSI EP17-A2 (June 2012)

Meaning
LOB Limit of blank
LOD Limit of detection
LLMI Lower limit of measuring interval (Limit of quantitation in earlier literature)

Task of this section

LOB, LOD, and LLMI should all be determined.

Materials and Time needed Tasks

3 days 60 replicates of blank sample
2 reagent lots 60 replicates of low concentration sample
4 blank samples (Bs) Repeat for each day
4 positive low concentration samples Repeat for each reagent lot
(LCs)
(at levels 1-5x LOB)
6 positive samples (spiked above LOD)

Calculation of LOB
Non-parametric (Preferred)
(1)Put all sample data in one row in an excel spreadsheet
(2)Sort the column (Data Sort)
(3)LOB = 0.5 * ([57th] + [58th])
If there are more than one reagent lot then use the largest value of all reagent lots

Parametric (Requirement: Gaussian distribution) assume data in A1:A60

(1)Calculate the mean of the samples (Excel: =average(A1:A60)) In cell B1
(2)Calculate the SD of the samples (Excel: =stdev(A1:A60)) In cell B2
(3)Assuming 4 blank sample used, 60 replicates used, then:
LOB = B1+B2*(1.645/(1-(1/(4*(60*4-4)))))

Calculation of LOD
Parametric analysis (Please perform transform if data is not normal) assume data in
C1:C60
(1)Calculate SD of low concentration data set (Excel: =stdev(C1:C60)) In cell D1
(2)Assuming 4 low concentration sample used, 60 replicates used, then:
LOD = LOB + D1*(1.645/(1-(1/(4*(60*4-4)))))
Demonstration (1 blank sample; 1 low concentration; 60 replicates)

Demonstration results

Calculation of LOQ / LLMI (Precision Profile Approach)

Standard deviation at different spiked concentration is plotted against concentration.
The simplest method is to choose the concentration tested, which has a coefficient of
variation below a certain percentage (say, 20%).

Alternatively, a linear or exponential model can be used. Demonstration below:

(1)Plot a function y = 0.2 * x (or any other desirable CV)
(2)Plot SD (y-axis) against spiked value (x-axis)
(3)Fit a general function y = (A1+A2*x)^A3 to the data in (2)

The desired LLMI is at the junction of the two lines.