Tag Archives: qPCR limit of detection

Data Analysis – RLOv DNA Helicase qPCR Limit of Detection Calcs

I previously ran three qPCR plates (20160121, 20160122, & 20160125) with 20 reps of each of the following copy numbers to determine the limit of detection (or, analytical sensitivity) of this qPCR assay: 30, 10, 3, 1

Additionally, I determined the average baseline threshold to set (580.5), based on eight qPCRs (see 20160128).

Performed calculations and determined limit of detection (defined as >95% of samples produced amplification) for this assay is three copies. The single copy sample amplified 88% of the time. This also means that our cycle threshold cut-off for this qPCR assay should be 37.9, as the mean Cq for three copies was 37.33, with a standard deviation of 0.53.

The data and calculations can be seen below (Google Sheet – scroll to the right to see calcs):

Google Sheet: RLOv_DNA_helicase_LoD_calcs

 

 

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qPCR – RLOv DNA Helicase Assay Limit of Detection

Continuing RLOv DNA Helicase qPCR assay validation.

This is the third of three plates to establish the assay’s limit of detection.

The first plate was run 20160121.

The second plate was run 20160122.

The limit of detection assessment is conducted in the following fashion:

  • Three plates of qPCRs; each plate run on different days.
  • On each plate; 20 reps each of the following standard curve copy numbers: 30, 10, 3, 1

Master mix calcs (Google Sheet): 201600121 – qPCR RLOv DNA Helicase Promega LoD-1

Plate layout, cycling params, etc can be seen in the qPCR Report (see Results below).

Results:
qPCR Report (PDF): Sam_2016-01-25 10-48-06_CC009827.pdf
qPCR Data File (CFX96): Sam_2016-01-25 10-48-06_CC009827.pcrd

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qPCR – RLOv DNA Helicase Assay Limit of Detection

Continuing RLOv DNA Helicase qPCR assay validation.

This is the second of three plates to establish the assay’s limit of detection. The first plate was run yesterday (20160121).

The limit of detection assessment is conducted in the following fashion:

  • Three plates of qPCRs; each plate run on different days.

  • On each plate; 20 reps each of the following standard curve copy numbers: 30, 10, 3, 1

Master mix calcs (Google Sheet): 201600121 – qPCR RLOv DNA Helicase Promega LoD-1

Plate layout, cycling params, etc can be seen in the qPCR Report (see Results below).

Results:
qPCR Report (PDF): Sam_2016-01-22 10-15-55_CC009827.pdf
qPCR Data File (CFX96): Sam_2016-01-22 10-15-55_CC009827.pcrd

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qPCR – RLOv DNA Helicase Assay Limit of Detection

Beginning RLOv DNA Helicase qPCR assay validation.

This is the first of three plates to establish the assay’s limit of detection.

The limit of detection assessment is conducted in the following fashion:

  • Three plates of qPCRs; each plate run on different days.

  • On each plate; 20 reps each of the following standard curve copy numbers: 30, 10, 3, 1

Master mix calcs (Google Sheet): 201600121 – qPCR RLOv DNA Helicase Promega LoD-1

Plate layout, cycling params, etc can be seen in the qPCR Report (see Results below).

Results:
qPCR Report (PDF): Sam_2016-01-21 15-12-31_CC009827.pdf
qPCR Data File (CFX96): Sam_2016-01-21 15-12-31_CC009827.pcrd

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qPCR – Promega 2x GoTaq Probe Master Mix Withering Syndrome Assay Limit of Detection

Ran limit of detection qPCR for the Promega 2x GoTaq Probe Master Mix using p18RK7 (from 20120730) primary curve and p18RK7 low curve (from 20140507).

Master mix calcs are here: 20140506 – qPCR WSN1 Promega LoD-1

Cycling params, plate layout, etc. can be found in the qPCR Report in the Results below.

Results:

qPCR Report (PDF): Sam_2014-05-21 14-04-07_CC009827.pdf
qPCR Data File (CFX96): Sam_2014-05-21 14-04-07_CC009827.pcrd

Data can be found here: Promega Master Mix Withering Syndrome Limit of Detection

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