Microfluidics and Organoids, the Power Couple of Developmental Biology and Oncology Studies.

Organoids are an advanced cell model that hold the key to unlocking a deeper understanding of in vivo cellular processes. This model can be used in understanding organ development, disease progression, and treatment efficacy. As the scientific world embraces the model, it must also establish the best practices for cultivating organoids and utilizing them to the greatest potential in assays. Microfluidic devices are emerging as a solution to overcome the challenges of organoids and adapt assays. Unfortunately, the various applications of organoids often depend on specific features in a device. In this review, we discuss the options and considerations for features and materials depending on the application and development of the organoid.

Enabling high-sensitivity live single-cell mass spectrometry using an integrated electrical lysis and nano electrospray ionization interface.

BACKGROUND: Live single-cell metabolomic studies encounter inherent difficulties attributed to the limited sample volume, minimal compound quantity, and insufficient sensitivity in the Mass Spectrometry (MS) method used to obtain single-cell data. However, understanding cellular heterogeneity, functional diversity, and metabolic processes within individual cells is essential. Exploring how individual cells respond to stimuli, including drugs, environmental changes, or signaling molecules, offers insights into biology, oncology, and drug discovery. Efficient release of cell contents (lysis) is vital for accurate metabolite detection at the single-cell level. Despite this, traditional approaches in live single cell metabolomics methods do not emphasize efficient lysis to prevent sample dilution. Instead, current live single cell metabolomics methods use direct infusion to introduce the cell into the mass spectrometry without prior chromatographic separation or a lysis step, which adversely affects sensitivity and metabolic coverage. RESULTS: To address this, we developed an integrated single-cell electrical lysis and nano spray (SCEL-nS) platform coupled to an Orbitrap MS capable of efficiently lysing a single cell after being sampled with specially manufactured micropipettes. Lysis efficiency was validated by comparing live cell stain fluorescent intensities of intact and electrically lysed cells through microscopy imaging. The SCEL-nS platform successfully induced the breakdown of a single cell, significantly reducing the live cell stain's fluorescent intensity indicating cell membrane breakdown. Additionally, SCEL-nS was validated by measuring single cells spiked with the anti-cancer drug tamoxifen by MS. SCEL-nS use resulted in statistically significant increase in the peak measured by the method compared to the traditional non-lysis method. SIGNIFICANCE: Overall, our results demonstrate that the newly incorporated SCEL-nS platform achieved higher sensitivities compared to traditional live single cell analysis methods.

Omicron infection-associated T- and B-cell immunity in antigen-naive and triple-COVID-19-vaccinated individuals.

Since early 2022, various Omicron variants have dominated the SARS-CoV-2 pandemic in most countries. All Omicron variants are B-cell immune escape variants, and antibodies induced by first-generation COVID-19 vaccines or by infection with earlier SARS-CoV-2 variants largely fail to protect individuals from Omicron infection. In the present study, we investigated the effect of Omicron infections in triple-vaccinated and in antigen-naive individuals. We show that Omicron breakthrough infections occurring 2-3.5 months after the third vaccination restore B-cell and T-cell immune responses to levels similar to or higher than those measured 14 days after the third vaccination, including the induction of Omicron-neutralizing antibodies. Antibody responses in breakthrough infection derived mostly from cross-reacting B cells, initially induced by vaccination, whereas Omicron infections in antigen-naive individuals primarily generated B cells binding to the Omicron but not the Wuhan spike protein. Although antigen-naive individuals mounted considerable T-cell responses after infection, B-cell responses were low, and neutralizing antibodies were frequently below the limit of detection. In summary, the detection of Omicron-associated B-cell responses in primed and in antigen-naive individuals supports the application of Omicron-adapted COVID-19 vaccines, but calls into question their suitability if they also contain/encode antigens of the original Wuhan virus.

BNT162b2-boosted immune responses six months after heterologous or homologous ChAdOx1nCoV-19/BNT162b2 vaccination against COVID-19.

Heterologous prime/boost vaccination with a vector-based approach (ChAdOx-1nCov-19, ChAd) followed by an mRNA vaccine (e.g. BNT162b2, BNT) has been reported to be superior in inducing protective immunity compared to repeated application of the same vaccine. However, data comparing immunity decline after homologous and heterologous vaccination as well as effects of a third vaccine application after heterologous ChAd/BNT vaccination are lacking. Here we show longitudinal monitoring of ChAd/ChAd (n = 41) and ChAd/BNT (n = 88) vaccinated individuals and the impact of a third vaccination with BNT. The third vaccination greatly augments waning anti-spike IgG but results in only moderate increase in spike-specific CD4 + and CD8 + T cell numbers in both groups, compared to cell frequencies already present after the second vaccination in the ChAd/BNT group. More importantly, the third vaccination efficiently restores neutralizing antibody responses against the Alpha, Beta, Gamma, and Delta variants of the virus, but neutralizing activity against the B.1.1.529 (Omicron) variant remains severely impaired. In summary, inferior SARS-CoV-2 specific immune responses following homologous ChAd/ChAd vaccination can be compensated by heterologous BNT vaccination, which might influence the choice of vaccine type for subsequent vaccination boosts.