Portable digital microfluidic serological immunoassays for measles and rubella were developed and evaluated in a remote setting.

We introduce Digital microfluidic Isolation of Single Cells for -Omics (DISCO), a platform that allows users to select particular cells of interest from a limited initial sample size and connects single-cell sequencing data to their immunofluorescence-based phenotypes. Specifically, DISCO combines digital microfluidics, laser cell lysis, and artificial intelligence-driven image processing to collect the contents of single cells from heterogeneous populations, followed by analysis of single-cell genomes and transcriptomes by next-generation sequencing, and proteomes by nanoflow liquid chromatography and tandem mass spectrometry. The results described herein confirm the utility of DISCO for sequencing at levels that are equivalent to or enhanced relative to the state of the art, capable of identifying features at the level of single nucleotide variations. The unique levels of selectivity, context, and accountability of DISCO suggest potential utility for deep analysis of any rare cell population with contextual dependencies.

Immunoprecipitation (IP) is a common method for isolating a targeted protein from a complex sample such as blood, serum, or cell lysate. In particular, IP is often used as the primary means of target purification for the analysis by mass spectrometry of novel biologically derived pharmaceuticals, with particular utility for the identification of molecules bound to a protein target. Unfortunately, IP is a labor-intensive technique, is difficult to perform in parallel, and has limited options for automation. Furthermore, the technique is typically limited to large sample volumes, making the application of IP cleanup to precious samples nearly impossible. In recognition of these challenges, we introduce a method for performing microscale IP using magnetic particles and digital microfluidics (DMF-IP). The new method allows for 80% recovery of model proteins from approximately microliter volumes of serum in a sample-to-answer run time of approximately 25 min. Uniquely, analytes are eluted from these small samples in a format compatible with direct analysis by mass spectrometry. To extend the technique to be useful for large samples, we also developed a macro-to-microscale interface called preconcentration using liquid intake by paper (P-CLIP). This technique allows for efficient analysis of samples >100× larger than are typically processed on microfluidic devices. As described herein, DMF-IP and P-CLIP-DMF-IP are rapid, automated, and multiplexed methods that have the potential to reduce the time and effort required for IP sample preparations with applications in the fields of pharmacy, biomarker discovery, and protein biology.

A pooled human serum sample with a haemagglutination-inhibition (HI) titre of 1/320 was used as the reference positive serum and a pooled human serum sample with no detectable antibodies to rubella-that is, HI titre < 1/10-was used as the reference negative serum. These sera, like all the test samples, were used at 1/100 dilution as determined by checkerboard titration as being optimal under the conditions of the test.Three hundred and sixty-seven single serum samples and 99 paired serum samples were tested for rubella antibodies by the HI test7 and retested by the micro-ELISA procedure similar to that described by Voller etal.2Amounts (200 ,ul) of1/100 dilution of serum samples were added to duplicate wells of the sensitised plates and these were incubated for two hours at room temperature. The plates were then shaken dry and washed in three changes of PBS-Tween. They were again shaken dry and to each well was added 200 ul of a 1/700 dilution of alkaline phosphatase conjugated antihuman globulin prepared in our own laboratory. After overnight incubation at 4°C, the plates were emptied and washed as before. A 200 ,ul amount of the substrate (p-nitrophenyl phosphate 1 mg/ml in 10% diethanolamine buffer, pH 9-8 with 0 5 mmol/l MgC12) was then added to each well. After 60 to 90 min, the reaction was stopped by the addition of 50 1I of 3 mol/l NaOH to each well. The time of stopping the reaction was based on the reference positive serum sample giving a reading of about 1-0 using a spectrophotometer to measure absorbance at 405 nm.