PCR Detection Kit

Definition

A dPCR detection kit is a commercial reagent kit specifically designed for digital PCR (DPCR).

Traditional PCR: 

DNA is exponentially amplified, and the copy number of the starting DNA is indirectly estimated using a fluorescent signal. This result is relative and susceptible to interference.

dPCR (digital PCR):

 Its core principle is "divide and conquer." A sample is divided into thousands of tiny droplets or reaction wells, each containing zero or a few target DNA molecules. Each droplet is then individually amplified by PCR. After amplification, the fluorescent signal from each droplet is detected, directly counting the number of droplets containing the target DNA (positive signal), thereby quantifying the exact copy number of the target DNA in the original sample without relying on a standard curve.

A dPCR detection kit is an all-in-one toolkit designed to accomplish this process. It typically contains:

1.Primers and probes: Core reagents that specifically recognize and amplify target gene sequences (such as viral RNA, cancer gene mutations, etc.).

2.PCR reaction mixture: Contains enzymes (reverse transcriptase, DNA polymerase), dNTPs, buffer, and all other chemical components required for PCR.

3.Droplet generator oil (for droplet-based dPCR): Used to partition the sample aqueous phase into thousands of nanoliter-scale droplets.

4.Positive control: Used to verify the success of the experiment.

5.Standard (provided in some kits): Used to verify quantitative accuracy.

Functions of dPCR Detection Kits

The core function of dPCR detection kits is to provide extremely sensitive and precise absolute quantification of trace amounts of specific nucleic acids (DNA or RNA). 

Specific features include:

1.Ultra-high sensitivity: Detects extremely low concentrations of target sequences, even at the single-molecule level, that are difficult to detect with conventional qPCR.

2.Absolute quantification: Directly provides absolute values such as "copies/µL" without the need for reference to a standard curve, resulting in more accurate and reliable results.

3.Strong anti-interference ability: High tolerance to PCR inhibitors (such as hemoglobin and heparin) present in the sample, as inhibitors are diluted into each microreaction unit and do not affect the overall situation.

4.Excellent Reproducibility: Because it is an endpoint test and direct enumeration, results are less variable between different laboratories and operators.

Disease-Related

Due to its superior performance, dPCR technology plays a vital role in the detection and research of a variety of diseases:

1.Cancer:

*Liquid biopsy: Detecting circulating tumor DNA (ctDNA) in the blood. ctDNA levels are extremely low, and dPCR can accurately detect rare gene mutations (such as EGFR, KRAS, and BRAF mutations) within ctDNA, enabling early cancer screening, medication guidance, efficacy monitoring, and recurrence warning.

*Minimal Residual Disease (MRD) Monitoring: Detecting the presence of minimal residual cancer cells after treatment to assess the risk of recurrence.

2.Infectious Diseases:

*Precise Viral Load Quantification: Accurate viral load monitoring of viruses such as HIV, HBV (Hepatitis B Virus), and HCV (Hepatitis C Virus) is crucial for evaluating the effectiveness of antiviral therapy.

*Emerging Infectious Diseases: During the COVID-19 pandemic, dPCR has been used as the "gold standard" for validating qPCR results, demonstrating significant advantages, particularly when testing samples with low viral loads (such as asymptomatic individuals) or resolving qPCR "gray zone" results.

3.Genetic Diseases:

*Rare Mutation Detection: Detects low-frequency mosaic mutations associated with genetic diseases.

*Copy Number Variant (CNV) Analysis: Accurately identifies gene copy number gains or losses (such as in Duchenne muscular dystrophy and spinal muscular atrophy), with greater accuracy than traditional methods.

4.Non-Invasive Prenatal Testing (NIPT):

Precisely assesses the risk of chromosomal aneuploidies (such as Down syndrome) by detecting cell-free fetal DNA in maternal peripheral blood.

5.Microbiology:

Absolute quantification of the abundance of specific bacterial species in complex samples such as environmental microbes and intestinal flora.

Who is Suitable for This Application

dPCR testing is generally not the first choice for routine screening, but rather serves specific populations:

1.Cancer Patients: Particularly those seeking targeted drug targets and those who need to monitor treatment efficacy and recurrence risk after treatment.

2.Patients with chronic infectious diseases, such as HIV and hepatitis B, require accurate viral load monitoring to adjust treatment plans.

3.High-risk individuals with a family history of genetic diseases require precise genetic counseling and carrier screening.

4.Pregnant women, especially those whose prenatal screening indicates a high risk and who require more accurate non-invasive prenatal testing.

5.Participants in clinical research: Patients in drug clinical trials who require accurate biomarker data for stratification.

Application Scenarios

The dPCR kit's application scenarios are primarily in scientific research and high-end clinical testing:

1.Clinical diagnostic laboratories:

*As a confirmation and supplemental tool for qPCR results, resolving difficult samples.

*Carrying out high-end projects such as tumor liquid biopsy, rare disease diagnosis, and precise viral load testing.

2.Biomedical R&D:

*Precise quantification of biomarkers in drug clinical trials.

*Quality control of cell and gene therapy products (e.g., vector copy number detection).

*Development and validation of companion diagnostic reagents.

3.Life science research:

*Gene expression analysis (especially for low-expressed genes).

*Verification of next-generation sequencing (NGS) results.

*Applications in fields requiring absolute quantification, such as environmental microbiology and food safety testing (e.g., quantification of genetically modified ingredients).

4.Forensic science:

Precise analysis of trace DNA samples.