A Multifunctional Frontloading Approach for Repeated Recycling of a Pressure-Controlled AFM Micropipette

Abstract

Fluid force microscopy combines the positional accuracy and force sensitivity of an atomic force microscope (AFM) with nanofluidics via a microchanneled cantilever. However, adequate loading and cleaning procedures for such AFM micropipettes are required for various application situations. Here, a new frontloading procedure is described for an AFM micropipette functioning as a force- and pressure-controlled microscale liquid dispenser. This frontloading procedure seems especially attractive when using target substances featuring high costs or low available amounts. Here, the AFM micropipette could be filled from the tip side with liquid from a previously applied droplet with a volume of only a few μL using a short low-pressure pulse. The liquid-loaded AFM micropipettes could be then applied for experiments in air or liquid environments. AFM micropipette frontloading was evaluated with the well-known organic fluorescent dye rhodamine 6G and the AlexaFluor647-labeled antibody goat anti-rat IgG as an example of a larger biological compound. After micropipette usage, specific cleaning procedures were tested. Furthermore, a storage method is described, at which the AFM micropipettes could be stored for a few hours up to several days without drying out or clogging of the microchannel. In summary, the rapid, versatile and cost-efficient frontloading and cleaning procedure for the repeated usage of a single AFM micropipette is beneficial for various application situations from specific surface modifications through to local manipulation of living cells, and provides a simplified and faster handling for already known experiments with fluid force microscopy.

Local tissue manipulation via a force- and pressure-controlled AFM micropipette for analysis of cellular processes

“…The micropipette was loaded with substances via the previously developed frontloading procedure 18 . In our present experiments, we used volumes of about 25 µL containing the target substance.…”

“…1 ). In this context, we apply the previously developed micropipette frontloading technique and the associated cleaning procedure 18 . Especially in physiological experiments, this method is very profitable because some substances, like the biogenic amine dopamine, are not stable for a long time in an aqueous environment.…”

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Figure 1 Schematic view of possible applications with the AFM micropipette at living tissue. Schematic illustration of ( a ) the previously published frontloading procedure 18 and ( b ) the loading position of the micropipette at living tissue with sufficient sealing between micropipette and tissue; estimated observation for ( c ) the CellTracker Red (CTR) and SYTO 9 Green local loading experiments and ( d ) the dopamine and sodium nitroprusside (SNP)-induced local manipulation experiments.

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