Use of high frequency electrorotation to identify cytoplasmic changes in cells non-disruptively.

In this paper we demonstrate how the use of frequencies ranging from 50 kHz to 5 GHz in the analysis of cells by electrorotation can open the path to the identification of differences not detectable by conventional set-ups. Earlier works usually reported electrorotation devices operating below 20 MHz, limiting the response obtained to properties associated with the cell membrane. Those devices are thus unable to resolve the physiological properties in the cytoplasm. We used microwave-based technology to extend the frequency operation to 5 GHz. At high frequencies (from tens of MHz to GHz), the electromagnetic signal passes through the membrane and allows probing the cytoplasm. This enables several applications, such as cell classification, and viability analysis. Additionally, the use of conventional microfabrication techniques reduces the cost and complexity of analysis, compared to other non-invasive methods. We demonstrated the potential of this set-up by identifying two different populations of T-lymphocytes not distinguishable through visual assessment. We also assessed the effect of calcein on cell cytoplasmic properties and used it as a controlled experiment to demonstrate the possibility of this method to detect changes happening predominantly in the cytoplasm.

Non-Invasive Microwave Hyperthermia and Simultaneous Temperature Monitoring with a Single Theranostic Applicator.

Cancer therapies are constantly evolving. Currently, heating tumor tissue is becoming more accessible as a stand-alone method or in combination with other therapies. Due to its multiple advantages over other heating mechanisms, microwave hyperthermia has recently gained a lot of traction. In this work, we present a complementary split-ring resonator that is simultaneously excited in two independent frequency bands. With a high-power signal, the applicator is excited and heats the tissue-under-test up to 50°C with an average heating rate of 0.72°C per second. Furthermore, we present a dielectric temperature control system using the same applicator for microwave hyperthermia applications, which currently still requires an additional thermometry system. By exciting the applicator with a low-power signal, we can constantly monitor its resonant frequency. This resonant frequency depends on the tissue properties, which in turn are temperature-dependent. In the temperature range from 20-50°C, a positive correlation between the temperature and resonant frequency was established.Clinical relevance - Exploiting the dual-band behavior of the complementary split-ring resonator to heat the tissue-under-test while dielectrically monitoring its temperature, creates new possibilities towards a theranostic, non-invasive microwave hyperthermia applicator.

Controlled Measurement Setup for Ultra-Wideband Dielectric Modeling of Muscle Tissue in 20-45 °C Temperature Range.

In order to design electromagnetic applicators for diagnostic and therapeutic applications, an adequate dielectric tissue model is required. In addition, tissue temperature will heavily influence the dielectric properties and the dielectric model should, thus, be extended to incorporate this temperature dependence. Thus, this work has a dual purpose. Given the influence of temperature, dehydration, and probe-to-tissue contact pressure on dielectric measurements, this work will initially present the first setup to actively control and monitor the temperature of the sample, the dehydration rate of the investigated sample, and the applied probe-to-tissue contact pressure. Secondly, this work measured the dielectric properties of porcine muscle in the 0.5-40 GHz frequency range for temperatures from 20 °C to 45 °C. Following measurements, a single-pole Cole-Cole model is presented, in which the five Cole-Cole parameters (ϵ∞, σs, Δϵ, τ, and α) are given by a first order polynomial as function of tissue temperature. The dielectric model closely agrees with the limited dielectric models known in literature for muscle tissue at 37 °C, which makes it suited for the design of in vivo applicators. Furthermore, the dielectric data at 41-45 °C is of great importance for the design of hyperthermia applicators.

Effect of Open-Ended Coaxial Probe-to-Tissue Contact Pressure on Dielectric Measurements.

Open-ended coaxial probes are widely used to gather dielectric properties of biological tissues. Due to the lack of an agreed data acquisition protocol, several environmental conditions can cause inaccuracies when comparing dielectric data. In this work, the effect of a different measurement probe-to-tissue contact pressure was monitored in the frequency range from 0.5 to 20 GHz. Therefore, we constructed a controlled lifting platform with an integrated pressure sensor to exert a constant pressure on the tissue sample during the dielectric measurement. In the pressure range from 7.74 kPa to 77.4 kPa, we observed a linear correlation of - 0 . 31 ± 0 . 09 % and - 0 . 32 ± 0 . 14 % per kPa for, respectively, the relative real and imaginary complex permittivity. These values are statistically significant compared with the reported measurement uncertainty. Following the literature in different biology-related disciplines regarding pressure-induced variability in measurements, we hypothesize that these changes originate from squeezing out the interstitial and extracellular fluid. This process locally increases the concentration of membranes, cellular organelles, and proteins in the sensed volume. Finally, we suggest moving towards a standardized probe-to-tissue contact pressure, since the literature has already demonstrated that reprobing at the same pressure can produce repeatable data within a 1% uncertainty interval.

Novel Fabrication Process for Integration of Microwave Sensors in Microfluidic Channels.

This paper presents a novel fabrication process that allows integration of polydimethylsiloxane (PDMS)-based microfluidic channels and metal electrodes on a wafer with a micrometer-range alignment accuracy. This high level of alignment accuracy enables integration of microwave and microfluidic technologies, and furthermore accurate microwave dielectric characterization of biological liquids and chemical compounds on a nanoliter scale. The microfluidic interface between the pump feed lines and the fluidic channels was obtained using magnets fluidic connection. The tube-channel interference and the fluidic channel-wafer adhesion was evaluated, and up to a pressure of 700 mBar no leakage was observed. The developed manufacturing process was tested on a design of a microwave-microfluidic capacitive sensor. An interdigital capacitor (IDC) and a microfluidic channel were manufactured with an alignment accuracy of 2.5 µm. The manufactured IDC sensor was used to demonstrate microwave dielectric sensing on deionized water and saline solutions with concentrations of 0.1, 0.5, 1, and 2.5 M.

A Microwave Platform for Reliable and Instant Interconnecting Combined with Microwave-Microfluidic Interdigital Capacitor Chips for Sensing Applications.

This work presents a novel platform conceived as an interconnect box (ICB) that brings high-frequency signals from microwave instruments to consumable lab-on-a-chip devices. The ICB can be connected to instruments with a standard coaxial connector and to consumable chips by introducing a spring-levered interface with elastomer conductive pins. With the spring-system, microwave-microfluidic chips can be mounted reliably on the setup in a couple of seconds. The high-frequency interface within the ICB is protected from the environment by an enclosure having a single slit for mounting the chip. The stability and repeatability of the contact between the ICB and inserted consumable chips are investigated to prove the reliability of the proposed ICB. Given the rapid interconnecting of chips using the proposed ICB, five different interdigital capacitor (IDC) designs having the same sensing area were investigated for dielectric permittivity extraction of liquids. The designed IDCs, embedded in a polydimethylsiloxane (PDMS) channel, were fabricated with a lift-off gold patterning technology on a quartz substrate. Water-Isopropanol (IPA) mixtures with different volume fractions were flushed through the channel over IDCs and sensed based on the measured reflection coefficients. Dielectric permittivity was extracted using permittivity extraction techniques, and fitted permittivity data shows good agreement with literature from 100 to 25 GHz.

An Interdigital Capacitor for Microwave Heating at 25 GHz and Wideband Dielectric Sensing of nL Volumes in Continuous Microfluidics.

This paper proposes a miniature microwave-microfluidic chip based on continuous microfluidics and a miniature interdigital capacitor (IDC). The novel chip consists of three individually accessible heaters, three platinum temperature sensors and two liquid cooling and mixing zones. The IDC is designed to achieve localized, fast and uniform heating of nanoliter volumes flowing through the microfluidic channel. The heating performance of the IDC located on the novel chip was evaluated using a fluorescent dye (Rhodamine B) diluted in demineralized water on a novel microwave-optical-fluidic (MOF) measurement setup. The MOF setup allows simultaneous microwave excitation of the IDC by means of a custom-made printed circuit board (connected to microwave equipment) placed in a top stage of a microscope, manipulation of liquid flowing through the channel located over the IDC with a pump and optical inspection of the same liquid flowing over the IDC using a fast camera, a light source and the microscope. The designed IDC brings a liquid volume of around 1.2 nL from room temperature to 100 °C in 21 ms with 1.58 W at 25 GHz. Next to the heating capability, the designed IDC can dielectrically sense the flowing liquid. Liquid sensing was evaluated on different concentration of water-isopropanol mixtures, and a reflection coefficient magnitude change of 6 dB was recorded around 8.1 GHz, while the minimum of the reflection coefficient magnitude shifted in the same frequency range for 60 MHz.