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Real-time PCR (qPCR) is a powerful technique for genotyping and gene expression analysis. Currently, qPCR experiments are becoming increasingly complex—involving an expansive and growing list of targets from a larger number of samples, all with more technical replicates. Accordingly, there is an increased need for high-throughput solutions to handle these challenges without sacrificing sensitivity, specificity, and reproducibility.
From our long history of developing innovative reagents for qPCR, we know the difficulties that can arise when trying to adapt complicated experiments to high-throughput workflows. The SmartChip Real-Time PCR System is a complete, high-throughput solution that enables an unrivaled level of assay and sample format flexibility.
The SmartChip Real-Time PCR System workflow is shown in Figure 1. SmartChip MyDesign Kits contain blank chips with 5,184 reaction nanowells and can be obtained as either single- or twenty-packs. The SmartChip Real-Time PCR System is composed of the SmartChip MultiSample NanoDispenser (MSND) and the SmartChip Real-Time PCR Cycler. The SmartChip MSND is a high-precision, nanoliter-volume liquid handling system capable of dispensing into 5,184 reaction nanowells in under 40 minutes. The SmartChip Cycler enables fast real-time analysis in two hours, supports probe- or dye-based assays such as FAM, VIC, ROX, and green intercalating dyes, and includes integrated analysis software that enables the interrogation of over 10,000 samples per day.
Unrivaled flexibility that delivers consistent results
The power of the SmartChip system is derived from the 5,184 individual nanowells on each chip provided in the SmartChip MyDesign Kit (Table I, Figure 2). The chips can be configured with 14 different assay and sample arrangements, allowing you to run the experiments you want to run instead of tailoring your experiments to fit rigid sample and assay formats.
Supported chip configurations
Table I. Range of assay and sample configurations with the SmartChip MyDesign Kit.
The SmartChip system utilizes a chip cooling station and enclosed, humidified environment to ensure thermal uniformity for high-precision results. A single assay run using the same sample in all 5,184 nanowells demonstrates consistent well-to-well performance with Ct values exhibiting a standard deviation <0.1. Thus, the chips enable reproducible and accurate results, with no artifacts or variability due to well location.
Efficiency without giving up sensitivity
The SmartChip system utilizes 100-nl reactions that offer significant reagent and cost savings over 25-μl reactions typically used in 384-well plates. 100-nl reactions, requiring only 3–10 ng/μl of input, are sensitive enough to eliminate the need for costly and time-consuming preamplification. Further reducing the reaction volume to 10 μl (as in an IFC format) provides only modest cost savings and requires 60 ng/μl of input, which does necessitate preamplification.
Highly reproducible gene expression measurements over a wide dynamic range
Accurate and sensitive detection of targets is critical for gene expression analysis. Using a titration of lambda DNA, we demonstrate that the SmartChip system is capable of detecting gene expression targets over a linear dynamic range greater than 6 logs, with a R2 value >0.99 (Figure 5). This means you can have confidence in your assay results.
It is essential that data generated is consistently reproducible, regardless of the number of days, operators, or replicates involved. Using the SmartChip system, three different users ran the same gene expression assay across two different chips on three different days. The data consistency is extremely high (R2 >0.98) from chip to chip, operator to operator, and day to day (Figure 6). Thus, you can expect consistent and reproducible performance from the SmartChip system, regardless of these variables.
The SmartChip system enables you to dispense assays and samples into blank chips or to dispense samples into preprinted chips. Moreover, you can easily switch between using blank and printed chips with no impact on your results. The same gene expression assay and dilution series were run on the SmartChip system using either blank chips from the SmartChip MyDesign Kit or custom printed chips. Linear correlation of Ct values was extremely high (R2 >0.99) and the standard deviation between experiments was <0.1 (Figure 7). Thus demonstrating that the SmartChip system can enable switching between gene discovery applications on blank chips and gene screening applications on printed chips without the need for revalidation.
Sensitive, accurate, and consistent genotyping
Accurate and sensitive detection is critical for making calls when genotyping. Samples of two different concentrations (1 ng/µl and 5 ng/µl) were run on the SmartChip system using the same genotyping assay (Figure 8). While the 1 ng/µl samples are more diffuse, the clustering still enables calling. Thus the SmartChip system demonstrates robust calling in samples as low as 1 ng/µl.
As with gene expression assays, the SmartChip system supports dispensing both genotyping assays and samples into blank chips, as well as dispensing only samples into preprinted chips. Critically, the data generated is consistent, regardless of the method used. The same genotyping assay was run on five different SmartChip systems using either blank chips from the SmartChip MyDesign Kit or custom printed chips (Figure 9). Both workflows produced high call rates with high concordance to known samples. Thus, the SmartChip system gives you the ability to run the experiments you want, the way you want, while delivering the accurate and consistent results you expect.
High-throughput real-time PCR
Where throughput meets flexibility
Real-time PCR (qPCR) is a powerful technique for genotyping and gene expression analysis. Currently, qPCR experiments are becoming increasingly complex—involving an expansive and growing list of targets from a larger number of samples, all with more technical replicates. The SmartChip Real-Time PCR System is a complete high-throughput solution that enables an unrivaled amount of flexible assay and sample formats, allowing researchers to seamlessly switch between dispensing assay reagents and samples into blank chips, or dispensing samples into custom, preprinted chips without the need for revalidation.