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Table 2 Comparison of seven assays for kdr genotyping based on specialist equipment required, cost, safety, simplicity of protocol and speed of method. Capital cost was calculated for all assays and is correct at the time of submission. Consumable/running cost was calculated for all assays except for the HOLA and SSOP-ELISA where the running cost listed was obtained from the original report of the method.

From: Detection of knockdown resistance (kdr) mutations in Anopheles gambiae: a comparison of two new high-throughput assays with existing methods

 

TaqMan

TaqMan Endpoint

HRM

AS-PCR

SSOP-ELISA

PCR Dot Blot

HOLA

Specialist equipment required

Real-time PCR machine

PCR thermocyler

Fluorimeter

High-spec real time PCR machine

PCR thermocycler

Gel electrophoresis and imaging equipment

PCR thermocycler

Shaking incubator

Multichannel pipette

*optional ELISA plate reader

PCR thermocycler

Shaking incubator

Multichannel pipette

PCR thermocycler

Multichannel pipette

*optional ELISA plate reader

Capital outlay cost (for equipment above given in US$)

96 well $25 000

48 well $19 000

$17 800

$50 000

$10 000

$7600

*optional ELISA plate reader add $5000

$7600

$5500

*optional ELISA plate reader add $5000

Hazardous chemicals

-

-

-

Ethidium bromide

TMAC, H2SO4, SDS

SDS

SDS

Protocol run time

1 hr 45 mins

2 hrs

1 hr 35 mins

~4 hrs 30 mins

~5 hrs 30 mins

~16–18 hrs

~6 hrs 30 mins

Number of steps

1

2

1

2

17

16

15

Primers/Probes required (for detection of 3 kdr alleles)

2 PCR primers

3 fluorescently labelled probes

2 PCR primers

3 fluorescently labelled probes

2 PCR primers

5 PCR primers

2 PCR primers (one biotin labelled)

3 SSOPs (digoxigenin labelled)

2 PCR primers

3 SSOPs (digoxigenin labelled)

2 PCR primers

2 reporter primers (5'phosphorylation and 3'fluorescein labelled)

4 detector primers (biotin labelled)

Number of tubes/wells/membranes required per sample

2

2

1

2

3

3

4

Running cost (per sample for three alleles)

$1.72

$1.72

$0.62

$0.92

sim $1

$1.6

$1.74