Existing methods for RNA diagnostics, such as reverse transcription PCR (RT-PCR), mainly rely on nucleic acid amplification (NAA) and RT processes, which are known to introduce substantial issues, including amplification bias, cross-contamination, and sample loss. To address these problems, we introduce a confinement effect-inspired Cas13a assay for single-molecule RNA diagnostics, eliminating the need for NAA and RT. This assay involves confining the RNAtriggered Cas13a catalysis system in cell-like-sized reactors to enhance local concentrations of target and reporter simultaneously, via droplet microfluidics. It achieves >10 000-fold enhancement in sensitivity when compared to the bulk Cas13a assay and enables absolute digital single-molecule RNA quantitation. We experimentally demonstrate its broad applicability for precisely counting microRNAs, 16S rRNAs, and SARS-CoV-2 RNA from synthetic sequences to clinical samples with excellent accuracy. Notably, this direct RNA diagnostic technology enables detecting a wide range of RNA molecules at the single-molecule level. Moreover, its simplicity, universality, and excellent quantification capability might render it to be a dominant rival to RT-qPCR.

In order to determine gene frequencies of human platelet antigen (HPA) and establish a panel of accredited HPA-1a, -2a, -4a, -5a and -6a-negative donors as well as an HPA-typed platelet donor registry, a total of 1000 Chinese donors of Han nationality (500 from north China and 500 from south China) were typed for HPA-1 through -16 using a DNA-based polymerase chain reaction with sequence-specific primers genotyping method. The gene frequencies of HPA-1b, -2b, -3b, -4b, -5b, -6bw, -10bw and -15b were 0.0060, 0.0485, 0.4055, 0.0045, 0.0140, 0.0135, 0.0005 and 0.4680, respectively. The HPA-7bw, -8bw, -9bw, -11bw, -12bw, -13bw, -14bw and -16bw alleles were not found. The HPA-2b and -5b homozygous donors were detected at low frequencies. The HPA mismatch probabilities potentially leading to alloimmunization in random platelet transfusion vary with a region from 0.1% to 37% depending on the distribution patterns of common and less common alleles in each system. This study provides a useful HPA-typed plateletpheresis donor registry in China and could improve platelet antibody detection and HPA-matched platelet transfusion in alloimmune thrombocytopenic patients.

This study provides the molecular bases of RHD alleles RHD 325del and RHD intron 2 1A. The existences of RHD 711del, RH (D1 CE2-9 D10), and RH (D1 CE2-9 D10) alleles in the Chinese population were confirmed. A PCR-SSP-based assay for rapid detection of RHD 325del and RHD intron 2 1A alleles was established and it could be used to predict the RHD genotype in the Chinese RhD-negative subjects.

BACKGROUND: This study aimed to analyze the mutation spectrum of the JK‐null phenotype in the Chinese population. The JK gene encoding the Kidd blood group antigen protein and JK*A/JK*B polymorphism caused by a G‐to‐A mutation at nt838 are well described. However, the molecular basis of the JK‐null phenotype in Chinese populations remains unclear. STUDY DESIGN AND METHODS: Sixteen unrelated JK‐null phenotype donors detected by red blood cell urea lysis resistance assay of 201,194 Chinese blood donors were confirmed in serologic agglutination tests. JK‐null alleles were analyzed by MnlI polymerase chain reaction–restriction fragment length polymorphism and sequencing of all JK gene coding regions. RESULTS: In addition to the well‐known Polynesian JK‐null allele JK*B(IVS5‐1g>a) and two alleles discovered in Taiwan, JK*B(896G>A) and JK*B(222C>A), seven JK‐null allele types were detected in this study including four novel JK‐null alleles: a nonsense mutation, JK*B(512G>A); two types of missense point mutations, JK*B(536C>G) and JK*B(437T>C); and a splice mutation, JK*A(IVS8+5g>a), resulting in skipping of Exon 8. CONCLUSION: This study demonstrates the frequency and heterogeneity of the JK‐null phenotype in Chinese populations. Based on our findings, the mechanisms underlying the Chinese Jk(a−b−) phenotype are quite different from other ethnic groups. The two most common types of JK‐null alleles were JK*B(IVS5‐1g>a) and JK*B(896G>A) in Chinese persons. Four novel JK‐null alleles were noted to be associated with the Jk(a−b−) phenotype.