Expanding the cell quantity of CRISPR/Cas9 gene editing by continuous microfluidic electroporation chip.

CRISPR/Cas9-mediated gene editing offers promising and safe therapeutic options for a wide range of diseases. The technical difficulty of efficiently acquiring large quantities of gene-edited therapeutic cells in a short time period is now preventing the widespread clinical application of CRISPR/Cas9-mediated gene editing. Herein, a Large Volume Continuous Electroporation Chip (LaViE-Chip) has been developed to address the challenge of acquiring sufficient quantities of genetically edited cells for CRISPR/Cas9 gene editing.

Quantitatively Evaluating Interactions between Patient-Derived Organoids and Autologous Immune Cells by Microfluidic Chip.

The coculture of patient-derived tumor organoids (PDOs) and autologous immune cells has been considered as a useful surrogate of tumor-immune environment. However, the immune interactions between PDOs and autologous immune cells, including immune-mediated killing behaviors and immune-related cytokine variations, have yet to be quantitatively evaluated. This study presents a microfluidic chip for quantifying interactions between PDOs and autologous immune cells (IOI-Chip).

A Microfluidic Chip-Based Automated System for Whole-Course Monitoring the Drug Responses of Organoids.

Tumor patients-derived organoids, as a promising preclinical prediction model, have been utilized to evaluate drug responses for formulating optimal therapeutic strategies. Detecting adenosine triphosphate (ATP) has been widely used in existing organoid-based drug response tests. However, all commercial ATP detection kits containing the cell lysis procedure can only be applied for single time point ATP detection, resulting in the neglect of dynamic ATP variations in living cells.

A dual-functional microfluidic chip for guiding personalized lung cancer medicine: combining EGFR mutation detection and organoid-based drug response test.

Many efforts have been paid to advance the effectiveness of personalized medicine for lung cancer patients. Sequencing-based molecular diagnosis of EGFR mutations has been widely used to guide the selection of anti-lung-cancer drugs. Organoid-based assays have also been developed to test individual responses to anti-lung-cancer drugs.

Rapid determination of the presence of EGFR mutations with DNA-based nanocalipers.

Selecting 1st-line treatment for lung cancer is currently a binary choice, either chemotherapy or targeted medicine, depending on whether EGFR mutations exist. Next-generation sequencing is fully capable of accurately identifying EGFR mutations and guiding the usage of tyrosine kinase inhibitors, but it is highly expensive. Moreover, as the sequencing is not helpful for patients with wild-type EGFR, the long wait for sequencing may delay the chemotherapy and correspondingly increase the risks of cancer progression.

Real-time monitoring ATP variation in human cancer organoids for a long term by DNA-based nanosensor.

Cancer organoids have become promising tools for predicting drug responses on many different types of cancer. Detecting the adenosine triphosphate (ATP) has currently been considered as a decisive test to profile the growth status and drug responses of organoids. ATP profiling using commercial ATP detection kits, which involve cell lysis, can be performed at a single time spot, causing a clinical dilemma of selecting the optimal time spot to adopt diverse cancer types and patients.

Mechanisms, Techniques and Devices of Airborne Virus Detection: A Review.

Airborne viruses, such as COVID-19, cause pandemics all over the world. Virus-containing particles produced by infected individuals are suspended in the air for extended periods, actually resulting in viral aerosols and the spread of infectious diseases. Aerosol collection and detection devices are essential for limiting the spread of airborne virus diseases.

Dual-function microneedle array for efficient photodynamic therapy with transdermal co-delivered light and photosensitizers.

Photodynamic therapy (PDT), as a globally accepted method for treating different forms of skin or mucosal disorders, requires efficient co-delivery of photosensitizers and corresponding therapeutic light. The adverse effects of intravenous injection of photosensitizers have been reduced by the development of microneedle arrays for transdermal local photosensitizer delivery. However, the drawbacks of the only available therapeutic light delivery method at the moment, which is directly applying light to the skin surface, are yet to be improved.

Methods and platforms for analysis of nucleic acids from single-cell based on microfluidics.

Single-cell nucleic acid analysis aims at discovering the genetic differences between individual cells which is well known as the cellular heterogeneity. This technology facilitates cancer diagnosis, stem cell research, immune system analysis, and other life science applications. The conventional platforms for single-cell nucleic acid analysis more rely on manual operation or bulky devices.

A microfluidic cytometer for white blood cell analysis.

Despite the wide use of cytometry for white blood cell classification, the performance of traditional cytometers in point-of-care testing remains to be improved. Microfluidic techniques have been shown with considerable potentials in the development of portable devices. Here we present a prototype of microfluidic cytometer which integrates a three-dimensional hydrodynamic focusing system and an on-chip optical system to count and classify white blood cells.

A Microfluidic Chip for Screening and Sequencing of Monoclonal Antibody at a Single-Cell Level.

The pairing of heavy and light chains of an antibody decides the specificity of monoclonal antibodies (mAbs). Acquisition of the genes encoding variable regions of paired heavy and light chains (V:V) is crucial, but it is a labor- and cost-intensive process in traditional methods. The emerging microfluidic chips have brought us to a portal of directly acquiring natively paired V:V genes by sequencing single target cells.

A Microfluidic Chip for Efficient Circulating Tumor Cells Enrichment, Screening, and Single-Cell RNA Sequencing.

Single-cell RNA sequencing on circulating tumor cells (CTCs) proves useful to study mechanisms of tumor heterogeneity, metastasis, and drug resistance. Currently, single-cell RNA sequencing of CTCs usually takes three prerequisite steps: enrichment of CTCs from whole blood, characterization of captured cells by immunostaining and microscopic imaging, and single-cell isolation through micromanipulation. However, multiple pipetting and transferring steps can easily cause the loss of rare CTCs.

A microfluidic chip for screening high-producing hybridomas at single cell level.

Hybridomas are a commonly used, or even the only option, for laboratory study and pilot production of monoclonal antibodies (mAbs), which are crucial for both targeted therapy and biomedical study. A long-term culture of hybridomas will inevitably induce a heterogenization of the whole hybridoma population, resulting in a continuous growth of non-producing hybridomas. To overcome the limits of existing methods of screening heterogeneous hybridomas, in which the whole multi-round screening process is performed in multi-well plates or other discrete modules, this study presents a novel method in which all processing steps of a multi-round hybridoma screening are finished in a single microfluidic chip.

Device for whole genome sequencing single circulating tumor cells from whole blood.

Whole-genome sequencing on circulating tumor cells (CTCs) at the single cell level has recently been found helpful for precision medicine, as the oncogenic profiles of single CTCs are useful for discovering oncogenic mutation heterogeneities and guiding/adjusting cancer treatment. To overcome the limits of existing methods of single CTC sequencing, in which CTC enrichment, identification and gene amplification are performed by discrete modules, this study presents a novel method in which all processing steps from blood sample collection to preparation of gene amplification products for sequencers are finished in a single microfluidic chip. This microfluidic chip comprehensively performs blood filtering, CTC enrichment, CTC identification/isolation, CTC lysis and whole genome amplification (WGA) at the single cell level.

An oligonucleotide synthesizer based on a microreactor chip and an inkjet printer.

Synthetic oligonucleotides (oligos) are important tools in the fields of molecular biology and genetic engineering. For applications requiring a large number of oligos with high concentration, it is critical to perform high throughput oligo synthesis and achieve high yield of each oligo. This study reports a microreactor chip for oligo synthesis.

Identifying EGFR-Expressed Cells and Detecting EGFR Multi-Mutations at Single-Cell Level by Microfluidic Chip.

EGFR mutations companion diagnostics have been proved to be crucial for the efficacy of tyrosine kinase inhibitor targeted cancer therapies. To uncover multiple mutations occurred in minority of EGFR-mutated cells, which may be covered by the noises from majority of un-mutated cells, is currently becoming an urgent clinical requirement. Here we present the validation of a microfluidic-chip-based method for detecting EGFR multi-mutations at single-cell level.

Combined peripheral natural killer cell and circulating tumor cell enumeration enhance prognostic efficiency in patients with metastatic triple-negative breast cancer.

Objective: Triple-negative breast cancer (TNBC) is a heterogeneous disease with poor prognosis. Circulating tumor cells (CTCs) are a promising predictor for breast cancer prognoses but their reliability regarding progression-free survival (PFS) is controversial. We aim to verify their predictive value in TNBC.

Quantitative Proteomic Analysis of Cell Responses to Electroporation, a Classical Gene Delivery Approach.

Electroporation, as an established nonviral technology for breaching cell membrane, has been accepted for the delivery of nucleic acids. Despite satisfactory delivery efficiencies have been achieved on multiple cell kinds by simply exhausting all possible electrical parameters, electroporation is still inefficient, or even invalid, for various kinds of cells. This is largely due to the lack of comprehensive understanding of cell responses to electrical stimulation at biological aspect.

Efficient delivery of nucleic acid molecules into skin by combined use of microneedle roller and flexible interdigitated electroporation array.

Delivery of nucleic acid molecules into skin remains a main obstacle for various types of gene therapy or vaccine applications. Here we propose a novel electroporation approach combined use of a microneedle roller and a flexible interdigitated electroporation array (FIEA) for efficient delivery of DNA and siRNA into mouse skin. Using micromachining technology, closely spaced gold electrodes were made on a pliable parylene substrate to form a patch-like electroporation array, which enabled close surface contact between the skin and electrodes.

Generation of a monoclonal antibody recognizing the heavily glycosylated CD45 protein and its application on identifying circulating tumor cells.

Here, we provide direct evidence that using recombinant proteins expressed in eukaryotic cells as antigen is a practical way to generate monoclonal antibodies (mAbs) against heavily glycosylated proteins. Heavily glycosylated proteins are typically difficult targets for mAb generation, being limited by unsatisfactory affinity and low specificity. Using the heavily glycosylated CD45 protein as an example, we demonstrate the entire process of expressing the protein in eukaryotic cells and using it as an antigen to generate CD45-targeting mAbs in mice.

Multiple single cell screening and DNA MDA amplification chip for oncogenic mutation profiling.

The oncogenic mutation heterogeneity of the cancer cell population has been proven to be essential for predicting both drug-response and drug-resistance of targeted therapies, such as tyrosine kinase inhibitors. It is necessary to accurately evaluate the mutation heterogeneity, oncogenic mutation and resistant mutation profiling at a single cell level. However, there are two major hurdles in the process.

Device To Study the Cell Invasion Behavior and Phenotypic Profile at Single Cell Level.

Multiple methods for investigating cell invasion behavior in vitro have proven useful in exploring the mechanisms behind the epithelial-mesenchymal transition (EMT) and EMT-related tumor cell invasion, for example, by revealing that cell heterogeneity existed in EMT. However, several hypotheses and predictions regarding EMT heterogeneity have remained unproven because of the inability to quantitatively profile cell invasion at the single cell level. Here, we present a microfluidic chip that provides the capability of simultaneously investigating single cell invasion behavior, phenotypic diversity, and responsiveness to anti-invasion drugs.