Controls for Real-Time PCR Infectious Disease Research Workflows

What are controls and why are they needed?

Controls play a vital role in helping to ensure the efficiency and accuracy of each stage within an infectious disease research workflow. By incorporating controls, we can closely monitor and evaluate the performance of the workflow as a whole. Controls enable assurance of reliability by validating the accuracy of the results obtained throughout the workflow, minimizing the risk of false-positive or false-negative outcomes.

Process controls:
- Used in the sample preparation and preamplification step to assess whether extraction and purification steps are effectively isolating and preserving nucleic acids of interest
- Used to evaluate the performance of preamplification

Amplification controls:
- Used in the PCR stage to help ensure reliability and accuracy of the assay

Overview of controls available for infectious disease research workflows

Process controls

Amplification controls

Extraction control

Spike-in controls

Amplification controls

Custom controls

Contains standardized samples that mimic, or are, assay targets
Employs a universal extraction control, Bacillus atrophaeus, a gram-positive bacteria that specially controls for cell lysis

Contains synthetic DNA sequences that act as an internal standard for subsequent workflow steps
3 spike-in controls available: RNA, DNA, and a control for panels containing antibiotic resistance targets

Controls for the amplification of the PCR step
Indicates assay is performing to standard
Available by application

Available to match the content of custom panel
Used to assess and confirm the amplification efficacy of selected assays during analytical validation, and as part of a routine workflow

Research workflow steps that involve the use of controls

Below, we’ve provided a process overview to demonstrate when to utilize process and amplification controls in a real-time PCR workflow for infectious disease research. While the specific example shown is for an OpenArray real-time PCR workflow, the steps are generally the same for TrueMark plates and cards.

Inventoried controls

Process controls for sample preparation and preamplification

TaqMan Universal Extraction Control Organism

TaqMan Spike in (Xeno) Controls

Run as separate sample
Goes into sample extraction, or it can be added directly to samples, which is optional
(B. atrophaeus), serves as a process control for cell lysis and nucleic acid recovery
The control is used with the proprietary TaqMan assay for Bacillus atrophaeus sequences

Spike into samples during and after sample extraction
Serves as a process control for the purification process and presence of inhibitors

Type of control	Catalog #	Product description	Volume
Extraction control	A39180	TaqMan Universal Extraction Control Organism	1 kit (3 pellets)
Spike-in control	A39175	TaqMan Universal DNA Spike-In Control	1,000 µL
Spike-in control	A39176	TaqMan Universal Universal DNA Spike-In Control (5X)	5 x 1,000 µL
Spike-in control	A50384	TaqMan Xeno Control, kanamycin resistance	1,000 uL
Spike-in / Reverse transcription control	A39179	TaqMan Universal RNA Spike In/Reverse Transcription (Xeno) Control	5 x 200 µL

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Amplification controls

Run amplification controls as a separate sample during the qPCR step to confirm assay performance at a known concentration. Amplification controls are available by application. If the application of interest is not listed in the table below, we also offer custom options to match the content of your panel.

Application	Catalog #	Product description	Volume
Vaginal microbiota	A32040	TaqMan Vaginal Microbiota Amplification Control	20 uL
Vaginal microbiota	A32913	TaqMan Vaginal Microbiota Amplification Control	250 uL
Urinary tract	A39174	TaqMan Urinary Tract Microbiota Amplification Control	5 x 50 uL
Respiratory tract	A39178	TaqMan Respiratory Tract Microbiota Amplification Control	5 x 50 uL
Respiratory tract	A48101	TrueMark Respiratory Panel 2.0 Amplification Control	250 uL
All	A50382	TrueMark Comprehensive Microbiota Control	1,000 uL

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Custom real-time PCR amplification controls

Synthetic amplification controls are available to match the exact content of your custom TrueMark real-time PCR panel. Our TaqMan Custom DNA Controls contain a linearized, multi-target plasmid pool used for microbial research and detection using TaqMan OpenArray Plates or TaqMan Array Cards.

Controls can be included as a positive control for panel-specific amplification
High concentration products (5 x 10⁷ copies/µL) can also be used to determine parameters necessary for analytical validation (e.g., limit of detection and Ct cut-off values)
During real-time PCR, the control can be used as a stand-alone sample to verify assay performance and help with troubleshooting

Catalog #	Product description	No. of targets	Concentration (copies/µL)	Volume (µL)
A50319	TrueMark Custom DNA Control	≤20	1 x 10⁵	1,000
A50375	TrueMark Custom DNA Control	21 to 30	1 x 10⁵	1,000
A50376	TrueMark Custom DNA Control	31 to 50	1 x 10⁵	1,000
A50377	TrueMark Custom DNA Control	>50	1 x 10⁵	1,000
A50378	TrueMark Custom DNA Control	≤20	5 x 10⁷	50
A50379	TrueMark Custom DNA Control	21 to 30	5 x 10⁷	50
A50380	TrueMark Custom DNA Control	31 to 50	5 x 10⁷	50
A50381	TrueMark Custom DNA Control	>50	5 x 10⁷	50

All TaqMan Custom DNA Control products should be stored at –15°C to –25°C. Please allow four weeks for your TaqMan Custom DNA Control product to be manufactured.

Ordering information

Ready to move forward with custom TrueMark real-time PCR solutions?

Complete this form to request a quote or consultation, or to place an order. A sales representative will reach out to walk you through the process of designing and implementing your new infectious disease research workflow.

For Research Use Only. Not for use in diagnostic procedures.