An integrated droplet digital polymerase chain reaction (ddPCR) gene chip that can quantify nucleic acid absolutely was constructed to avoid problems such as droplet fusion, fragmentation and sample contamination during pipetting. The integrated ddPCR gene chip was fabricated by photolithography and cyclic olefin copolymer (COC) injection mold. The 20 μL sample was distributed to about 60,000 water-in-oil droplets with a diameter of about 87 μm within 3 min by flow focus microstructure, and the volume of each droplet was about 0.27 nL. The generation of bubbles was reduced by reducing the width of the collection chamber and increasing the deceleration step. In addition, the efficiency of PCR was improved using COC materials. The sample of human epidermal growth factor receptor (EGFR) exon18 gene was quantitatively detected by this chip. Experimental results show that the detection signal has a good linear relationship with the DNA concentration from 10¹ to 10⁵ copies/μL (R² = 0.999) so that the integrated ddPCR gene chip can realize absolute quantification of nucleic acid. It is verified that the integration of droplet generation, PCR amplification and fluorescence detection are realized on one chip, reduce bubble generation and improve PCR efficiency. Compared with the current independent split chip, the integrated ddPCR gene chip ensures a high degree of anti-pollution performance, avoids the operation of droplet movement, reduces human interference, and standardizes the results. It has wide application prospects in nucleic acid quantification and trace detection.

构建了可绝对定量核酸的集成液滴数字聚合酶链反应（ddPCR）基因芯片，避免了移液过程中液滴融合、碎裂和样品污染等问题。集成的ddPCR基因芯片通过光刻和环烯烃共聚物（COC）注塑模具制造。20 μL样品通过流动聚焦微结构在3 min内分布到约60,000个直径约87 μm的油包水液滴中，每个液滴的体积约为0.27 nL。通过减小收集室的宽度和增加减速步骤来减少气泡的产生。此外，使用 COC 材料提高了 PCR 的效率。该芯片对人表皮生长因子受体（EGFR）外显子18基因样本进行定量检测。¹到10 ^{5 个}拷贝/μL（R ²  = 0.999），使得集成的ddPCR基因芯片可以实现核酸的绝对定量。经验证，在一个芯片上实现了液滴生成、PCR扩增和荧光检测的集成，减少了气泡的产生，提高了PCR效率。与目前的独立分体芯片相比，集成的ddPCR基因芯片保证了高度的抗污染性能，避免了飞沫运动的操作，减少了人为干扰，结果标准化。在核酸定量和痕量检测方面具有广泛的应用前景。