UMBC Center for Advanced Sensor Technology (CAST)

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The Center for Advanced Sensor Technology promotes the development of optical based sensors for biomedical, bioprocess, environmental and homeland defense applications. We are a multidisciplinary group of chemists, molecular biologists, electrical engineers and chemical and biochemical engineers. Our main objective is to create innovative sensor technology that is high throughput and low-cost amenable for transfer to industry counterparts.

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    Recent Developments in Bioprocess Monitoring Systems
    (Springer, 2024-01-11) Rahmatnejad, Vida; Wei, Yunqian; Rao, Govind
    Bioprocess monitoring systems are essential tools for achieving an optimal manufacturing process for pharmaceuticals. In recent years, there have been advancements in the development of these systems. One of the most notable developments in bioprocess monitoring systems is the use of biosensors, which can detect and measure biological molecules such as enzymes and proteins. The use of imaging technologies, such as microscopy and flow cytometry, has allowed for the non-invasive monitoring of cell culture, providing valuable information for process optimization. Other non-invasive monitoring systems are being developed with the aim of reducing the risk of contamination. Another important advancement is the use of machine learning and artificial intelligence in bioprocess monitoring systems. These technologies have the ability to learn from historical data and predict future outcomes, which can be beneficial for optimizing bioprocesses in real-time. Overall, recent advancements in sensor technology can help in improving the efficiency and effectiveness of bioprocessing. The integration of advanced sensing technologies, and machine learning can provide valuable insights and improve the ability to monitor and control bioprocesses online. In this chapter, we will discuss recent advancements in bioprocess monitoring systems, specifically for monoclonal antibody and cell and gene therapy manufacturing processes.
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    Global nuclear radiation monitoring using plants
    (SPIE, 2015-05-13) Islam, Mohammad; Romero-Talamás, Carlos; Kostov, Dan; Wang, Wanpeng; Liu, Zhongchi; Hussey, Daniel S.; Baltic, Eli; Jacobson, David L.; Gu, Jerry; Choa, Fow-Sen
    Plants exhibit complex responses to changes in environmental conditions such as radiant heat flux, water quality, airborne pollutants, soil contents. We seek to utilize the natural chemical and electrophysiological response of plants to develop novel plant-based sensor networks. Our present work focuses on plant responses to high-energy radiation – with the goal of monitoring natural plant responses for use as benchmarks for detection and dosimetry. For our study, we selected a plants cactus, Arabidopsis, Dwarf mango (pine), Euymus and Azela. We demonstrated that the ratio of Chlorophyll a to Chlorophyll b of the leaves has changed due to the exposure gradually come back to the normal stage after the radiation die. We used blue laser-induced blue fluorescence-emission spectra to characterize the pigment status of the trees. Upon blue laser excitation (400 nm) leaves show a fluorescence emission in the red spectral region between 650 and 800nm (chlorophyll fluorescence with maxima near 690nm and 735 nm). Sample tree subjects were placed at a distance of 1m from NIST-certified 241AmBe neutron source (30 mCi), capable of producing a neutron field of about 13 mrem/h. This corresponds to an actual absorbed dose of ~ 1 mrad/h. Our results shows that all plants are sensitive to nuclear radiation and some take longer time to recover and take less. We can use their characteristics to do differential detection and extract nuclear activity information out of measurement results avoid false alarms produced environmental changes. Certainly the ultimate verification can be obtained from genetic information, which only need to be done when we have seen noticeable changes on plant optical spectra, mechanical strength and electrical characteristics.
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    Rapid Bacterial Detection and Identification of Bacterial Strains Using Machine Learning Methods Integrated With a Portable Multichannel Fluorometer
    (IEEE, 2023-08-09) Hasan, Md Sadique; Sundberg, Chad; Hasan, Hasibul; Kostov, Yordan; Ge, Xudong; Choa, Fow-Sen; Rao, Govind
    Rapid and sensitive bioburden detection is of paramount importance in different applications including public health, and food and water safety. To overcome the traditional limitations of bacterial detection i.e., lengthy culture time, and complicated procedure, a low-cost, portable multichannel fluorometer coupled with machine learning (ML) has been implemented in this study. Five different strains of bacterial samples were tested along with the negative control for time-series fluorescence data collection and analysis. We applied different conventional unsupervised and supervised machine learning techniques with extracted features followed by preprocessing of the data. Initially, machine learning algorithms were applied for the qualitative detection of bacteria by binary classification followed by regression analysis to predict the level of contamination for E. coli. The multiclass classification was used to identify gram-positive, and gram-negative bacterial strains and differentiate all the bacterial strains tested. Our results show that around 97.9% accuracy can be achieved for bacterial contamination detection for as low as 1 CFU/mL while 92.1% accuracy can be achieved for differentiating the gram-positive and gram-negative strains. Additionally, with 1 minute of data, high accuracy is obtained for detecting bioburden, proving the multichannel fluorometer’s rapid detection capability. The multichannel fluorometer integrated with ML analytics is capable of automating data analysis and determining accurate and rapid bacterial detection on-site with the prediction of bioburden levels and differentiating bacterial strains and the protocol can be applied to the biosensors with a similar data type.
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    Tuning of antimicrobial property of commercial drugs by use of bio- synthesized noble metal nanoparticles
    (2014-02) Srinivasan, Venkatesh; Vedantam, Pallavi; Ramkumar, C. B.; Ramamurthy, Sai Sathish
    Silver nanoparticles are known to have antimicrobial and antifungal properties. Here we explore the effect of biogenic silver (AgNPs) and gold nanoparticles (AuNPs) on the antimicrobial property of drugs namely Vancomycin, Ampicillin and Tetracyclin, by measuring their zone of inhibition in disk diffusion studies against Escherichia coli, Enterococcus faecalis and Staphylococcus aureus. The different AuNPs and AgNPs used were synthesized using aqueous extracts of floral parts of Tecoma stans, Caesalpinia sappan, Tagetus erecta. The bacterial strains were spread on agar plates and the disks impregnated with known concentration of drug were used for disk diffusion studies and well diffusion studies. 20nm & 200nm commercially available AuNPs and AgNPs were also used along with drugs for disk diffusion studies and the results were compared. Different nanoparticles had different effect on drug's antimicrobial activity. The range of tunability of antimicrobial activity of drug achieved by using the biogenic nanoparticles was more than that of commercial nanoparticles. E.coli and S.aureus are known to gain resistance against Ampicilin and Vancomysin respectively. In this work, the effect of Ampicillin on E.coli and the effect of Vancomysin on S.aureus could be restored by using it along with our nanoparticles. Thus we propose a safe and simple method to increase or decrease drug's activity against specific microbe.
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    Ultra-Amplification of Surface Plasmon Coupled Emission Using an Engineered Graphene-Silver Thin Film Hybrid
    (Optica, 2012-12-12) Mulpur, Pradyumna; Vemu, Sai Krishna; Lingam, Kiran; Srinivasan, Venkatesh; Ramamurthy, Sai Sathish; Kamisetti, Venkataramaniah; Rao, Apparao M
    We report the selective ultra-amplification of the Surface Plasmon Coupled Emission by synergistic plasmon coupling on incorporating a novel hybrid thin film stacking of graphene on silver.
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    Novel Synthesis of Nanoparticles for Enhancements in Surface Plasmon Coupled Emission
    (Optica, 2014-12-16) Srinivasan, Venkatesh; Srikanth, M.; Nayak, Lisa; Ramamurthy, Sai Sathish; Neeleshwar, S.
    We report a novel synthesis route for nanomaterials namely SnSe and SnTe and their application as a simple thin film spacers for achieving 30–45 fold fluorescence enhancements using Surface Plasmon Coupled Emission platform.
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    Spacer layer engineering for ultrasensitive Hg(II) detection on surface plasmon-coupled emission platform
    (De Gruyter, 2017-06-15) Badiya, Pradeep Kumar; Jayakumar, Tejkiran Pindi; Srinivasan, Venkatesh; Ramamurthy, Sai Sathish
    In this work, we demonstrate for the first time the ultrasensitive detection of Hg²⁺ ions with femtomolar sensitivity in water samples with the use of the surface plasmon-coupled emission (SPCE) platform. The use of portable network diagnostic tools for water security and integrated water shed management is a topic of recent research interest. In this context, the current study explores Hg²⁺ monitoring using a rhodamine-6G (Rh6G) derivative bearing a monothiospirolactone mounted onto a SPCE substrate. Thus far, the limit of detection for mercury ions by the conventional fluorescence technique has been 0.15 nm. However, we have achieved 1 fm Hg²⁺ detection using silver nanoparticle-based spacer layer engineering on an SPCE sensor chip. Using this technology, a field device can be fabricated for rapid, ultrasensitive, multi-analyte detection (of contaminants) in water samples.
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    An approach to rapid distributed manufacturing of broad spectrum anti-viral griffithsin using cell-free systems to mitigate pandemics
    (2022-12-20) Borhani, Shayan G.; Levine, Max Z.; Krumpe, Lauren H.; Wilson, Jennifer; Henrich, Curtis J.; O’Keefe, Barry R.; Lo, Donald; Sittampalam, G. Sitta; Godfrey, Alexander G.; Lunsford, R. Dwayne; Mangalampalli, Venkata; Tao, Dingyin; LeClair, Christopher A.; Thole, Aaron; Frey, Douglas; Swartz, James; Rao, Govind
    This study describes the cell-free biomanufacturing of a broad-spectrum antiviral protein, griffithsin (GRFT) such that it can be produced with consistent purity and potency in less than 24 hours. We demonstrate GRFT production using two independent cell-free systems, one plant and one microbial. Griffithsin purity and quality were verified using standard regulatory metrics. Efficacy was demonstrated in vitro against SARS-CoV-2 and HIV-1 and was nearly identical to that of GRFT expressed in vivo. The proposed production process is efficient and can be readily scaled up and deployed anywhere in the world where a viral pathogen might emerge. The current emergence of viral variants has resulted in frequent updating of existing vaccines and loss of efficacy for front-line monoclonal antibody therapies. Proteins such as GRFT with its efficacious and broad virus neutralizing capability provide a compelling pandemic mitigation strategy to promptly suppress viral emergence at the source of an outbreak.
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    Microwave induced thermally assisted solvent-based bonding of biodegradable thermoplastics: an eco-friendly rapid approach for fabrication of microfluidic devices and analyte detection
    (Nature, 2022-09-27) Hasan, Md Sadique; Borhani, Shayan; Ramamurthy, Sai Sathish; Andar, Abhay; Ge, Xudong; Choa, Fow-Sen; Kostov, Yordan; Rao, Govind
    There is an increasing interest in low-cost, facile and versatile thermoplastic bonding for microfluidic applications that can be easily transitioned from laboratory prototyping to industrial manufacturing. In addition, owing to the surge in the usage of thermoplastic microfluidics and its adverse effect on the environment, it is prudent to source alternative materials that are biodegradable, providing a sustainable, green approach. To address the problems, here we introduce an environment friendly, low-cost and safe welding technology used in the fabrication of microcassettes from biodegradable cellulose acetate (CA) thermoplastics. The thermally assisted solvent based bonding of the thermoplastics was accomplished in a domestic microwave oven with the aid of a polyether ether ketone (PEEK) vise. To characterize the quality of the bonding, our in-house technique was compared with a conventional thermally assisted solvent bonding configuration using a heat press machine and tested under different conditions. Our microwave induced bonding of CA presents three times reduced bonding time with higher bonding strength, good reliability and does not necessitate the use of cumbersome instrumentation. Finally, we demonstrate an electrophoresis application and vitamin C detection accomplished using this biodegradable microcassette presenting comparable results with traditional techniques, illustrating the potential of this fabrication technique in different microfluidic applications.
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    Rapid ultra-sensitive and high-throughput bioburden detection: Microfluidics and instrumentation
    (ACS, 2022-06-06) Hasan, Md Sadique; Marsafari, Monireh; Tolosa, Michael; Andar, Abhay; Ramamurthy, Sai Sathish; Ge, Xudong; Kostov, Yordan; Rao, Govind
    Contamination detection often requires lengthy culturing steps to detect low-level bioburden. To increase the rate of detection and decrease the limit of detection (LOD), a system featuring microfluidics and a multichannel fluorometer has been developed. The eight-channel fluorometer enables parallel testing of multiple samples with the LOD as low as <1 cfu/mL. This low-cost system utilizes the slope of fluorescence intensity that serves as the criterion for bioburden detection. The redox indicator dye resazurin is used to monitor the presence of viable cells in this study and is reduced to resorufin with a high quantum yield at 585 nm. The sample under investigation is spiked with resazurin and loaded in a special-design microfluidic cassette, and the rate of change is observed via the fluorometer. The method was validated using primary Escherichia coli culture in comparison with a spectrophotometer which served as the gold standard. An optimized assay based on Luria–Bertani medium was developed. The impact on the assay sensitivity based on incubation and filtration steps was also explored. The assay is shown to pick up inadvertent contamination from test tubes and pipette tips showing its applicability in real-world settings. The data analysis demonstrated a comparable performance of the multichannel fluorometer vis-a-vis the conventional plate reader. The multichannel system is shown to detect bioburden presence in as low as 20 s for bacterial concentrations ≥5 cfu/mL after 6 h of incubation. Considering its portability, low cost, simplicity of operation, and relevant assay sensitivity, the system is well positioned to detect low-level bioburden in the laboratory, pharmaceutical, and field settings.
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    Disposable low-cost cardboard incubator for thermoregulation of stable preterm infant – a randomized controlled non-inferiority trial
    (Elsevier, 2020-12-07) Chandrasekaran, Ashok; Amboiram, Prakash; Balakrishnan, Umamaheswari; Abiramalatha, Thangaraj; Rao, Govind; Jan, Shaik Mohammad Shafi; Rajendran, Usha Devi; Sekar, Uma; Thiruvengadam, Gayathri; Ninan, Binu
    Background Incubators and radiant warmers are essential equipment in neonatal care, but the typical 1,500 to 35,000 USD cost per device makes it unaffordable for many units in low and middle-income countries. We aimed to determine whether stable preterm infants could maintain thermoregulation for 48 h in a low-cost incubator (LCI). Methods The LCI was constructed using a servo-heater costing 200 USD and cardboard infant-chamber. We conducted this open-labeled non-inferiority randomized controlled trial in a tertiary level teaching hospital in India from May 2017 to March 2018. Preterm infants on full feeds and receiving incubator or radiant warmer care were enrolled at 32 to 36 weeks post-menstrual age. We enrolled 96 infants in two strata (Strata-1< 33 weeks, Strata-2 ≥ 33 weeks at birth). Infants were randomized to LCI or standard single-wall incubator (SSI) after negative incubator cultures and monitored for 48 h in air-mode along with kangaroo mother care. The incubator temperature was adjusted manually to maintain skin and axillary temperatures between 36.5 °C and 37.5 °C. During post-infant period after 48 h, SSI and LCI worked for 5 days and incubator temperatures were measured. The primary outcome was maintenance of skin and axillary temperatures with a non-inferiority margin of 0.2 °C. Failed thermoregulation was defined as abnormal axillary temperature (< 36.5 °C or >37.5 °C) for > 30 continuous-minutes. Secondary outcomes were incidence of hypothermia and required incubator temperature. Trial registration details: Clinical Trial Registry - India (CTRI/2015/10/006316). Findings Prior to enrollment 79(82%) infants were in radiant warmer and 17(18%) infants were in incubator care. Median weight at enrollment in Strata-1 and Strata-2 for SSI vs. LCI was 1355(IQR 1250–1468) vs. 1415(IQR 1280–1582) and 1993(IQR 1595–2160) vs. 1995(IQR 1632–2237) grams. Mean skin temperature in Strata-1 and Strata-2 for SSI vs. LCI was 36.8 °C ± 0.2 vs. 36.7 °C ± 0.18 and 36.8 °C ± 0.22 vs. 36.7 °C ± 0.19. Mean axillary temperature in Strata-1 and Strata-2 for SSI vs. LCI was 36.9 °C ± 0.19 vs. 36.8 °C ± 0.16 and 36.8 °C ± 0.2 vs. 36.8 °C ± 0.19. Mixed-effect model done for repeated measures of skin and axillary temperatures showed the estimates were within the non-inferiority limit; -0.07 °C (95% CI -0.11 to -0.04) and -0.06 °C (95% CI -0.095 to -0.02), respectively. Failed thermoregulation did not occur in any infants. Mild hypothermia occurred in 11 of 48(23%) of SSI and 16 of 48(33%) of LCI, OR 1.28 (95%CI 0.85 to 1.91). Incubator temperature in LCI was higher by 0.7 °C (95%CI 0.52 to 0.91). In the post-infant period SSI and LCI had excellent reliability to maintain set-temperature with intra-class correlation coefficient of 0.93 (95%CI 0.92 to 0.94) and 0.96 (95%CI 0.96 to 0.97), respectively. Interpretation Maintenance of skin and axillary temperature of stable preterm infants in LCI along with kangaroo mother care was non-inferior to SSI, but at a higher incubator temperature by 0.7 °C. No adverse events occurred and LCI had excellent reliability to maintained set-temperature.
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    Developments in Insulin Manufacturing Using Cell-Free Systems
    Borhani, Shayan; Gurramkonda, Chandrasekhar; Frey, Douglas; Tolosa, Leah; Swartz, James; Rao, Govind
    The primary aim of this method is to emulate the naturally occurring mechanisms which take place in vivo, when converting preproinsulin into metabolically active insulin (4). Figure 2, displays the binding of pre-proinsulin to a CM HiTrap FF column (pH 3.0) followed by cleavage and threonine addition to the terminal B29 residue. The final product is then eluted using a linear salt gradient.
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    Robust and easy macrofluidic connections in acrylic
    (Royal Society of Chemistry, 2012-04-23) Horner, Nathalie; Henderson, Robert; Selock, Nick; Rao, Dr. Govind
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    Sensor Techknowledge: Sensors for Process Development: A Practical Guide
    (UMBC Center for Advanced Sensor Technology and Department of Biochemical and Environmental Engineering, 2020-07-05) Rao, Govind
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    Real‐time dissolved carbon dioxide monitoring II: Surface aeration intensification for efficient CO2 removal in shake flasks and mini‐bioreactors leads to superior growth and recombinant protein yields
    (Wiley Online Library, 2019-12-13) Chopda, Viki R.; Holzberg, Timothy; Ge, Xudong; Folio, Brandon; Wong, Lynn; Tolosa, Michael; Kostov, Yordan; Tolosa, Leah; Rao, Govind
    Mass transfer is known to play a critical role in bioprocess performance and henceforth monitoring dissolved O₂ (DO) and dissolved CO₂ (dCO₂) is of paramount importance. At bioreactor level these parameters can be monitored online and can be controlled by sparging air/oxygen or stirrer speed. However, traditional small‐scale systems such as shake flasks lack real time monitoring and also employ only surface aeration with additional diffusion limitations imposed by the culture plug. Here we present implementation of intensifying surface aeration by sparging air in the headspace of the reaction vessel and real‐time monitoring of DO and dCO₂ in the bioprocesses to evaluate the impact of intensified surface aeration. We observed that sparging air in the headspace allowed us to keep dCO₂ at low level, which significantly improved not only biomass growth but also protein yield. We expect that implementing such controlled smart shake flasks can minimize the process development gap which currently exists in shake flask level and bioreactor level results.
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    Real‐time dissolved carbon dioxide monitoring I: Application of a novel in situ sensor for CO2 monitoring and control
    (Wiley Online Library, 2019-12-16) Chopda, Viki R.; Holzberg, Timothy; Ge, Xudong; Folio, Brandon; Tolosa, Michael; Kostov, Yordan; Tolosa, Leah; Rao, Govind
    Dissolved carbon dioxide (dCO₂) is a well‐known critical parameter in bioprocesses due to its significant impact on cell metabolism and on product quality attributes.Processes run at small‐scale faces many challenges due to limited options for modular sensors for online monitoring and control. Traditional sensors are bulky, costly, and invasive in nature and do not fit in small‐scale systems. In this study, we present the implementation of a novel, rate‐based technique for real‐time monitoring of dCO₂ in bioprocesses. A silicone sampling probe that allows the diffusion of CO₂ through its wall was inserted inside a shake flask/bioreactor and then flushed with air to remove the CO₂ that had diffused into the probe from the culture broth (sensor was calibrated using air as zero‐point calibration). The gas inside the probe was then allowed to recirculate through gas‐impermeable tubing to a CO₂ monitor. We have shown that by measuring the initial diffusion rate of CO₂ into the sampling probe we were able to determine the partial pressure of the dCO₂ in the culture. This technique can be readily automated, and measurements can be made in minutes.Demonstration experiments conducted with baker’s yeast and Yarrowia lipolytica yeast cells in both shake flasks and mini bioreactors showed that it can monitor dCO₂ in real‐time. Using the proposed sensor, we successfully implemented a dCO₂‐based control scheme, which resulted in significant improvement in process performance.
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    Minimally invasive technique for measuring transdermal glucose with a fluorescent biosensor
    (Springer Nature Switzerland AG., 2018-08-31) Brown, Sheniqua; Zambrana, Paige N.; Ge, Xudong; Bagdure, Dayanand; Stinchcomb, Audra L.; Rao, Govind; Tolosa, Leah
    There is a need for blood glucose monitoring techniques that eliminate the painful and invasive nature of current methods, while maintaining the reliability and accuracy of established medical technology. This research aims to ultimately address these shortcomings in critically ill pediatric patients. Presented in this work is an alternative, minimally invasive technique that uses microneedles (MN) for the collection of transdermal glucose (TG). Due to their comparable skin properties, diffusion studies were performed on full thickness Yucatan miniature pig skin mounted to an in-line diffusion flow cell and on different skin sites of human subjects. Collected TG samples were measured with a L255C mutant of the E. coli glucose-binding protein (GBP) with an attached fluorescent probe. The binding constant (Kd = 0.67 μM) revealed the micromolar sensitivity and high selectivity of the his-tagged GBP biosensor for glucose, making it suitable for TG measurements. In both the animal and human models, skin permeability and TG diffusion across the skin increased with MN application. For intact and MN-treated human skin, a significant positive linear correlation (r > 0.95, p < 0.01) existed between TG and BG. The micromolar sensitivity of GBP minimized the volume required for interstitial fluid glucose analysis allowing MN application time (30 s) to be shortened compared to other studies. This time reduction can help in eliminating skin irritation issues and improving practical use of the technique by caregivers in the hospital. In addition, the his-tagged optical biosensor used in this work can be immobilized and used with a portable sensing fluorometer device at the point of care (POC) making this minimally invasive technology more ideal for use in the pediatric intensive care unit.
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    Minimally invasive technique for measuring transdermal glucose with a fluorescent biosensor
    (Springer Berlin Heidelberg, 2018-08-17) Brown, Sheniqua; Zambrana, Paige N.; Ge, Xudong; Bagdure, Dayanand; Stinchcomb, Audra L.; Rao, Govind; Tolosa, Leah
    There is a need for blood glucose monitoring techniques that eliminate the painful and invasive nature of current methods, while maintaining the reliability and accuracy of established medical technology. This research aims to ultimately address these shortcomings in critically ill pediatric patients. Presented in this work is an alternative, minimally invasive technique that uses microneedles (MN) for the collection of transdermal glucose (TG). Due to their comparable skin properties, diffusion studies were performed on full thickness Yucatan miniature pig skin mounted to an in-line diffusion flow cell and on different skin sites of human subjects. Collected TG samples were measured with a L255C mutant of the E. coli glucose-binding protein (GBP) with an attached fluorescent probe. The binding constant (Kd = 0.67 μM) revealed the micromolar sensitivity and high selectivity of the his-tagged GBP biosensor for glucose, making it suitable for TG measurements. In both the animal and human models, skin permeability and TG diffusion across the skin increased with MN application. For intact and MN-treated human skin, a significant positive linear correlation (r > 0.95, p < 0.01) existed between TG and BG. The micromolar sensitivity of GBP minimized the volume required for interstitial fluid glucose analysis allowing MN application time (30 s) to be shortened compared to other studies. This time reduction can help in eliminating skin irritation issues and improving practical use of the technique by caregivers in the hospital. In addition, the his-tagged optical biosensor used in this work can be immobilized and used with a portable sensing fluorometer device at the point of care (POC) making this minimally invasive technology more ideal for use in the pediatric intensive care unit.
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    Improving the recombinant human erythropoietin glycosylationusing microsome supplementation in CHO cell-free system
    (Wiley, 2018) Gurramkonda, Chandrasekhar; Rao, Aniruddha; Borhani, Shayan; Pilli, Manohar; Deldari, Sevda; Ge, Xudong; Pezeshk, Niloufar; Tolosi, Michael; Kostov, Yordon; Tolosa, Leah; Frey, Douglas D.; Rao, Govind
    Cell-Free Protein Synthesis (CFPS) offers many advantages for the production of recombinant therapeutic proteins using the HO cell-free system. However, many complex proteins are still difficult to express using this method. To investigate the current bottlenecks in cell-free glycoprotein production, we chose erythropoietin (40% glycosylated), an essential endogenous hormone which stimulates the development of red blood cells. Here, we report the production of recombinant erythropoietin (EPO) using CHO cell-free system. Using this method, EPO was expressed and purified with a twofold increase in yield when the cell-free reaction was supplemented with CHO microsomes. The protein was purified to near homogeneity using an ion-metal affinity column. We were able to analyze the expressed and purified products (glycosylated cell-free EPO runs at 25–28 kDa, and unglycosylated protein runs at 20 kDa on an SDS–PAGE), identifying the presence of glycan moieties by PNGase shift assay. The purified protein was predicted to have ∼2,300 IU in vitro activity. Additionally, we tested the presence and absence of sugars on the cell-free EPO using a lectin-based assay system. The results obtained in this study indicate that microsomes augmented in vitro production of the glycoprotein is useful for the rapid production of single doses of a therapeutic glycoprotein drug and to rapidly screen glycoprotein constructs in the development of these types of drugs. CFPS is useful for implementing a lectin-based method for rapid screening and detection of glycan moieties, which is a critical quality attribute in the industrial production of therapeutic glycoproteins.