Grafting of Maleic Anhydride onto Poly(vinylidene fluoride) Using Reactive Extrusion.

Poly(vinylidene fluoride) was grafted with maleic anhydride through reactive extrusion by using diisopropyl benzene peroxide as an initiator and 9-vinyl anthracene as a stabilizer. Effects of various parameters on grafting degree were investigated including the amounts of monomer, initiator and stabilizer. The maximum extent of grafting achieved was 0.74%. The graft polymers were characterized using FTIR, water contact angle, thermal, mechanical and XRD studies. Improved hydrophilic and mechanical properties were observed for graft polymers.

Single pixel hyperspectral Cherenkov-excited fluorescence imaging with LINAC X-ray sheet scanning and spectral unmixing.

Cherenkov light induced from megavolt (MV) X-rays during external beam radiotherapy serves as an internal light source to excite phosphors or fluorophores within biological tissues for molecular imaging. The broad spectrum of Cherenkov light leads to significant spectral overlap with any luminescence emission and, to overcome this problem, a single pixel hyperspectral imaging methodology was demonstrated here by coupling the detection with light sheet scanning and filtered back projection reconstruction of hyperspectral images. Thin scanned sheets of MV X-rays produce Cherenkov light to illuminate the planes deep within the tissue-simulating media. A fluorescence probe was excited by Cherenkov light, and a complete hyperspectral sinogram of the data was obtained through translation and rotation of the beam. Hyperspectral 2D images finally were reconstructed. Through this approach of spectral unmixing, it was possible to resolve hyperspectral images of both the Cherenkov and resulting fluorescence intensity from molecular sensors.

Time-gated luminescence imaging for background free in vivo tracking of single circulating tumor cells.

Fluorescence imaging is severely limited by the background and autofluorescence of tissues for in vivo detection of circulating tumor cells (CTCs). Time-gated luminescence (TGL) imaging, in combination with luminescent probes that possess hundreds of microsecond emission lifetimes, can be used to effectively suppress this background, which has predominantly nanosecond lifetimes. This Letter demonstrates the feasibility of TGL imaging using luminescent probes for the in vivo real time imaging and tracking of single CTCs circulating freely in the blood vessels with higher accuracy given by substantially higher signal-to-noise ratio. The luminescent probe used in this Letter was a commercial Eu3+ chelate (EuC) nanosphere with a super-long lifetime of near 800 µs, which enabled TGL imaging to achieve background-free detection with ∼5 times higher SNR versus steady state. Phantom and in vivo mouse studies indicated that EuC labeled tumor cells moving in medium or bloodstream at the speed of 1-2 mm/s could be captured in real time.

Establishment of a duplex SYBR green I-based real-time polymerase chain reaction assay for the rapid detection of canine circovirus and canine astrovirus.

The similar clinical characteristics of canine circovirus (CaCV) and canine astrovirus (CaAstV) infections and high frequency of co-infection make diagnosis difficult. In this study, a duplex SYBR Green I-based real-time polymerase chain reaction (PCR) assay was established for the rapid, simultaneous detection of CaCV and CaAstV. Two pairs of specific primers were designed based on the Rep gene of CaCV and the Cap gene of CaAstV. By using the real-time PCR assay method, the two viruses can be distinguished by the difference in melting temperatures, 79 °C and 86 °C for CaCV and CaAstV, respectively. This assay had high specificity, showing no cross-reaction with other common canine viruses, as well as high sensitivity, with minimum detection limits of 9.25 × 101 copies/µL and 6.15 × 101 copies/µL for CaCV and CaAstV, respectively. Based on the mean coefficient of variation, the method had good reproducibility and reliability. In a clinical test of 57 fecal samples, the rates of positive detection by real-time PCR were 14.04% (8/57) and 12.28% (7/57) for CaCV and CaAstV, respectively, and the rate of co-infection was 8.77% (5/57). In conclusion, the newly established duplex SYBR Green I-based real-time PCR assay is sensitive, specific, reliable, and rapid and is an effective tool for the detection of co-infections with CaCV and CaAstV.

Simultaneous detection of duck circovirus and novel goose parvovirus via SYBR green I-based duplex real-time polymerase chain reaction analysis.

Beak atrophy and dwarfism syndrome (BADS) is commonly caused by co-infection with duck circovirus (DuCV) and novel goose parvovirus (NGPV). Therefore, concurrent detection of both viruses is important for monitoring and limiting BADS, although such a diagnostic test has not been reported. In this study, we developed a duplex, SYBR Green I-based real-time polymerase chain reaction (PCR) assay to enable the simultaneous detection of DuCV and NGPV. The assay readily distinguished between the two viruses, based on their different melting temperatures (Tm), where the Tm for DuCV was 80 °C and that for NGPV was 84.5 °C. Other non-target duck viruses that were tested did not show melting peaks. The detection limit of the duplex assay was 101 copies/µL for both viruses. This method exhibited high repeatability and reproducibility, and both the inter-assay and intra-assay variation coefficients were <1.6%. Thirty-one fecal samples were collected for clinical testing using real-time PCR analysis, and the results were confirmed using sequencing. The rate of co-infection was 6.5%, which was consistent with the sequencing results. This duplex real-time PCR assay offers advantages over other tests, such as rapid, sensitive, specific, and reliable detection of both viruses in a single sample, which enables the quantitative detection of DuCV and NGPV in clinical samples. Using this test may be instrumental in reducing the incidence of BADS and the associated economic losses in the duck and goose industries.

TaqMan-based real-time polymerase chain reaction assay for specific detection of bocavirus-1 in domestic cats.

Feline bocavirus-1 (FBoV-1) was first discovered in Hong Kong in 2012, and studies have indicated that the virus may cause feline hemorrhagic enteritis. Currently, there is a lack of an effective and quantitative method for FBoV-1 detection. In this study, a TaqMan-based quantitative real-time PCR (qPCR) for FBoV-1 detection was established. Primers and probes were designed to target the conserved region of the FBoV-1 NS1 gene. The sensitivity analysis indicated that the minimum detection limit was 4.57 × 101 copies/µL. The specificity test revealed no cross-reaction with seven other common feline viruses, including the same species-FBoV-2 and FBoV-3. The sensitivity of this method was 100 times higher than that of conventional PCR (cPCR). The established method showed good repeatability, with the intra-assay and inter-assay coefficients of variation of 0.18%-1.00% and 0.27%-0.45%, respectively. Furthermore, the analysis of feline feces revealed that the detection rate by qPCR was 7.0% (9/128), whereas that by cPCR was 4.7% (6/128). In conclusion, the established qPCR assay can quantitatively detect FBoV-1 with a high sensitivity, high specificity, and good reproducibility, making it a promising technique for the clinical detection of and basic and epidemiological research on FBoV-1.

Identification and full-genome sequencing of canine kobuvirus in canine fecal samples collected from Anhui Province, eastern China.

Canine kobuvirus (CaKoV), a newly described virus, is the causative agent of gastroenteritis in dogs. In this study, 57 fecal samples from dogs with diarrhea in Anhui Province, eastern China, were collected. Among these, five samples were identified to be infected with CaKoV, by polymerase chain reaction targeting the CaKoV 3D gene. The five CaKoV strains were subjected to phylogenetic analysis. The sequences of VP1 from the five CaKoV strains were 93.6%-96.1% identical to each other and 91.75%-97.95% identical to other reported CaKoV VP1 sequences. In addition, the complete genome of one strain was successfully amplified and sequenced. The genome consisted of 8223 nucleotides and shared 94.6%-97.0% nucleotide and 93.1%-94.0% amino acid sequence identity with other CaKoV isolates. Phylogenetic analysis revealed that the CaKoV strain from Anhui Province was similar to other Chinese strains, and it was more closely related to feline and mouse kobuviruses than to sheep and bovine kobuviruses. Interestingly, all of the CaKoV-positive samples were coinfected with canine parvovirus. The finding of CaKoV infection in dogs with diarrhea and coinfection with canine parvovirus are a cause for concern and highlight the need for management and preventive measures.

Tomographic Cherenkov-excited luminescence scanned imaging with multiple pinhole beams recovered via back-projection reconstruction.

Cherenkov-excited luminescence scanned imaging (CELSI) is achieved with a clinical linear accelerator during external beam radiotherapy to map out molecular luminescence intensity or lifetime in tissue. In order to realize a deeper imaging depth with a reasonable spatial resolution in CELSI, we optimized the original scanning procedure to complete this in a way similar to x-ray computed tomography and with image reconstruction from maximum-likelihood expectation maximization and multi-pinhole irradiation for parallelization. Resolution phantom studies showed that a 0.3 mm diameter capillary tube containing 0.01 nM luminescent nanospheres could be recognized at a depth of 21 mm into tissue-like media. Small animal imaging with a 1 mm diameter cylindrical target demonstrated that fast 3D data acquisition can be achieved by this multi-pinhole collimator approach to image high-resolution luminescence through a whole animal.

Observation of short wavelength infrared (SWIR) Cherenkov emission.

Cherenkov emission induced by external beam radiation from a clinical linear accelerator has been shown in preclinical molecular imaging and clinical imaging. The broad spectrum Cherenkov emission should have a short wavelength infrared (SWIR, 1000-1700 nm) component, as predicted theoretically. To the best of our knowledge, this Letter is the first experimental observation of this SWIR Cherenkov emission induced by external beam radiation. The measured spectrum of SWIR Cherenkov emission matches the theoretical prediction, with a fluence rate near one-third of the visible and near-infrared red emissions (Vis-NIR, 400-900 nm). Imaging in water-based phantoms and biological tissues indicates that there is a sufficient fluence rate for radiotherapy dosimetry applications. The spatial resolution is improved approximately 5.3 times with SWIR Cherenkov emission detection versus Vis-NIR Cherenkov emission, which provides some improvement in the potential for higher resolution Cherenkov emission dosimetry and molecular sensing during clinical radiotherapy by imaging with SWIR wavelengths.

Addition of T2-guided optical tomography improves noncontrast breast magnetic resonance imaging diagnosis.

BACKGROUND: While dynamic contrast-enhanced magnetic resonance imaging (DCE MRI) is recognized as the most sensitive examination for breast cancer detection, it has a substantial false positive rate and gadolinium (Gd) contrast agents are not universally well tolerated. As a result, alternatives to diagnosing breast cancer based on endogenous contrast are of growing interest. In this study, endogenous near-infrared spectral tomography (NIRST) guided by T2 MRI was evaluated to explore whether the combined imaging modality, which does not require contrast injection or involve ionizing radiation, can achieve acceptable diagnostic performance. METHODS: Twenty-four subjects-16 with pathologically confirmed malignancy and 8 with benign abnormalities-were simultaneously imaged with MRI and NIRST prior to definitive pathological diagnosis. MRIs were evaluated independently by three breast radiologists blinded to the pathological results. Optical image reconstructions were constrained by grayscale values in the T2 MRI. MRI and NIRST images were used, alone and in combination, to estimate the diagnostic performance of the data. Outcomes were compared to DCE results. RESULTS: Sensitivity, specificity, accuracy, and area under the curve (AUC) of noncontrast MRI when combined with T2-guided NIRST were 94%, 100%, 96%, and 0.95, respectively, whereas these values were 94%, 63%, 88%, and 0.81 for DCE MRI alone, and 88%, 88%, 88%, and 0.94 when DCE-guided NIRST was added. CONCLUSION: In this study, the overall accuracy of imaging diagnosis improved to 96% when T2-guided NIRST was added to noncontrast MRI alone, relative to 88% for DCE MRI, suggesting that similar or better diagnostic accuracy can be achieved without requiring a contrast agent.

Characteristics and rehabilitation for patients with spinal cord stab injury.

[Purpose] The objective of the study was to compare the incidence, diagnosis, treatment, and prognosis of patients with spinal cord stab injury to those with the more common spinal cord contusion injury. [Subjects] Of patients hospitalized in China Rehabilitation Research Center from 1994 to 2014, 40 of those having a spinal cord stab injury and 50 with spinal cord contusion were selected. [Methods] The data of all patients were analyzed retrospectively. The cases were evaluated by collecting admission and discharge ASIA (American Spinal Injury Association) and ADL (activity of daily living) scores. [Results] After a comprehensive rehabilitation program, ASIA and ADL scores of patients having both spinal cord stab injury and spinal cord contusion significantly increase. However, the increases were noted to be higher in patients having a spinal cord stab injury than those having spinal cord contusion. [Conclusion] Comprehensive rehabilitation is effective both for patients having spinal cord stab injury and those with spinal cord contusion injury. However, the prognosis of patients having spinal cord stab injury is better than that of patients with spinal cord contusion.

Patient-specific arterial input function for accurate perfusion assessment in intraoperative fluorescence imaging.

Significance: The arterial input function (AIF) plays a crucial role in correcting the time-dependent concentration of the contrast agent within the arterial system, accounting for variations in agent injection parameters (speed, timing, etc.) across patients. Understanding the significance of the AIF can enhance the accuracy of tissue vascular perfusion assessment through indocyanine green-based dynamic contrast-enhanced fluorescence imaging (DCE-FI). Aim: We evaluate the impact of the AIF on perfusion assessment through DCE-FI. Approach: A total of 144 AIFs were acquired from 110 patients using a pulse dye densitometer. Simulation and patient intraoperative imaging were conducted to validate the significance of AIF for perfusion assessment based on kinetic parameters extracted from fluorescence images before and after AIF correction. The kinetic model accuracy was evaluated by assessing the variability of kinetic parameters using individual AIF versus population-based AIF. Results: Individual AIF can reduce the variability in kinetic parameters, and population-based AIF can potentially replace individual AIF for estimating wash-out rate ( k ep ), maximum intensity ( I max ), ingress slope with lower differences compared with those in estimating blood flow, volume transfer constant ( K trans ), and time to peak. Conclusions: Individual AIF can provide the most accurate perfusion assessment compared with assessment without AIF or based on population-based AIF correction.

Onward to Better Surgery - the Critical Need for Improved Ex Vivo Testing and Training Methods.

Guided surgery has demonstrated significant improvements in patient outcomes in some disease processes. Interest in this field has led to substantial growth in the technologies under investigation. Most likely no single technology will prove to be "best," and combinations of macro- and microscale guidance-using radiological imaging navigation, probes (activatable, perfusion, and molecular-targeted; large- and small-molecule), autofluorescence, tissue intrinsic optical properties, bioimpedance, and other characteristics-will offer patients and surgeons the greatest opportunity for high-success/low-morbidity medical interventions. Problems are arising, however, from the lack of valid testing formats; surgical training simulators suffer the same problems. Small animal models do not accurately recreate human anatomy, especially in terms of tissue volume. Large animal models are expensive and have difficulty replicating many pathological states, particularly when molecular specificity for individual cancers is required. Furthermore, the sheer number of technologies and the potential for synergistic combination leads to exponential growth of testing requirements that is unrealistic for in vivo testing. Therefore, critical need exists to expand the ex vivo/in vitro testing platforms available to investigators and, once validated, a need to increase the acceptance of these methods for funding and regulatory endpoints. Herein is a review of the available ex vivo/in vitro testing formats for guided surgery, a review of their advantages/disadvantages, and consideration for how our field may safely and more swiftly move forward through stronger adoption of these testing and validation methods.

Stable tissue-mimicking phantoms for longitudinal multimodality imaging studies that incorporate optical, CT, and MRI contrast.

Significance: Tissue phantoms that mimic the optical and radiologic properties of human or animal tissue play an important role in the development, characterization, and evaluation of imaging systems. Phantoms that are easily produced and stable for longitudinal studies are highly desirable. Aim: A new type of long-lasting phantom was developed with commercially available materials and was assessed for fabrication ease, stability, and optical property control. Magnetic resonance imaging (MRI) and x-ray computed tomography (CT) contrast properties were also evaluated. Approach: A systematic investigation of relationships between concentrations of skin-like pigments and composite optical properties was conducted to realize optical property phantoms in the red and near-infrared (NIR) wavelength range that also offered contrast for CT and MRI. Results: Phantom fabrication time was < 1 h and did not involve any heating or cooling processes. Changes in optical properties were < 2 % over a 12-month period. Phantom optical and spectral features were similar to human soft tissue over the red to NIR wavelength ranges. Pigments used in the study also had CT and MRI contrasts for multimodality imaging studies. Conclusions: The phantoms described here mimic optical properties of soft tissue and are suitable for multimodality imaging studies involving CT or MRI without adding secondary contrast agents.

Monitoring optoporated process on mammalian cells by real-time measurement of membrane resealing time.

Significance: Resealing time based loading efficiency of optoporation is the key parameter for drug or gene delivery. This work describes a comparatively simple optical approach to directly measure the cell membrane resealing time of the gold nanoparticle mediated photoporation. Aim: To establish a membrane potential detection optical system, which can provide a direct measurement of resealing time of the optoporated cells. Approach: Voltage sensitive dye has been used to label the gold nanoparticle covered cell before laser activation and the resealing time was estimated from the voltage change due to the fluorescence light intensity change before and after laser activation. The approach has been validated by the simulated data based on diffusion model and Monte Carlo simulation and the experimental data obtained from a flow cytometry analysis. Results: The measured resealing time after perforation varied from 28.6 to 163.8 s on Hela cells when the irradiation fluence was increased, with a correlation coefficient (R2) of 0.9938. This result is in agreement with the resealing time (1-2 min) of photothermal porated Hela cells measured by electrical impedance method. The intracellular delivery efficiency of extracellular macromolecular under the same irradiation fluence depends mainly on diffusion velocity rather than pore size. Conclusion: The method described here can be used to directly measure resealing time of optoporated cells for accurately estimating the loading efficiency on discovering the mechanism of optoporation.

Near-Infrared Spectral Tomography for Predicting Residual Cancer Burden during Early-Stage Neoadjuvant Chemotherapy for Breast Cancer.

PURPOSE: The aim of this study is to investigate whether near-infrared spectral tomography (NIRST) might serve as a reliable prognostic tool to predict residual cancer burden (RCB) in patients with breast cancer undergoing neoadjuvant chemotherapy (NAC) based upon early treatment response measurements. EXPERIMENTAL DESIGN: A total of thirty-five patients with breast cancer receiving NAC were included in this study. NIRST imaging was performed at multiple time points, including: before treatment, at end of the first cycle, at the mid-point, and post-NAC treatments. From reconstructed NIRST images, average values of total hemoglobin (HbT) were obtained for both the tumor region and contralateral breast at each time point. RCB scores/classes were assessed by a pathologist using histologic slides of the surgical specimen obtained after completing NAC. Logistic regression of the normalized early percentage change of HbT in the tumor region (ΔHbT%) was used to predict RCB and determine its significance as an indicator for differentiating cases within each RCB class. RESULTS: The ΔHbT% at the end of the first cycle, as compared with pretreatment levels, showed excellent prognostic capability in differentiating RCB-0 from RCB-I/II/III or RCB-II from RCB-0/I/III (P < 0.001). Corresponding area under the curve (AUC) values for these comparisons were 0.97 and 0.94, and accuracy values were 0.90 and 0.83, respectively. CONCLUSIONS: NIRST holds promise as a potential clinical tool that can be seamlessly integrated into existing clinical workflow within the infusion suite. By providing early assessment of RCB, NIRST has potential to improve breast cancer patient management strategies.

Fluorescence-guided and molecularly-guided debridement: identifying devitalized and infected tissue in orthopaedic trauma.

Following orthopaedic trauma, bone devitalization is a critical determinant of complications such as infection or nonunion. Intraoperative assessment of bone perfusion has thus far been limited. Furthermore, treatment failure for infected fractures is unreasonably high, owing to the propensity of biofilm to form and become entrenched in poorly vascularized bone. Fluorescence-guided surgery and molecularly-guided surgery could be used to evaluate the viability of bone and soft tissue and detect the presence of planktonic and biofilm-forming bacteria. This proceedings paper discusses the motivation behind developing this technology and our most recent preclinical and clinical results.

Proceduralist criteria for evaluating interface utility of novel imaging modalities in early phase clinical trials: evaluating the need for standardized criteria.

Accelerating innovation in the space of fluorescence imaging for surgical applications has increased interest in safely and expediently advancing these technologies to clinic through Food and Drug Administration-(FDA-) compliant trials. Conventional metrics for early phase trials include drug safety, tolerability, dosing, and pharmacokinetics. Most procedural imaging technologies rely on administration of an exogenous fluorophore and concurrent use of an imaging system; both of which must receive FDA approval to proceed to clinic. Because fluorophores are classified as medical imaging agents, criteria for establishing dose are different, and arguably more complicated, than therapeutic drugs. Since no therapeutic effect is desired, medical imaging agents are ideally administered at the lowest dose that achieves adequate target differentiation. Because procedural imaging modalities are intended to enhance and/or ease proceduralists' identification or assessment of tissues, beneficial effects of these technologies may manifest in the form of qualitative endpoints such as: 1) confidence; 2) decision-making; and 3) satisfaction with the specified procedure. Due to the rapid expansion of medical imaging technologies, we believe that our field requires standardized criteria to evaluate existing and emerging technologies objectively so that both quantitative and qualitative aspects of their use may be measured and useful comparisons to assess their relative value may occur. Here, we present a 15-item consensus-based survey instrument to assess the utility of novel imaging technologies from the proceduralist's standpoint.

Automated motion artifact correction for dynamic contrast-enhanced fluorescence imaging during open orthopedic surgery.

Indocyanine green (ICG)-based dynamic contrast-enhanced fluorescence imaging (DCE-FI) can objectively assess bone perfusion intraoperatively. However, it is susceptible to motion artifacts due to patient's involuntary respiration during the 4.5-minute DCE-FI data acquisition. An automated motion correction approach based on mutual information (MI) frameby-frame was developed to overcome this problem. In this approach, MIs were calculated between the reference and the adjacent frame translated and the maximal MI corresponded to the optimal translation. The images obtained from eighteen amputation cases were utilized to validate the approach and the results show that this correction can significantly reduce the motion artifacts and can improve the accuracy of bone perfusion assessment.

Spatial and temporal patterns in dynamic-contrast enhanced intraoperative fluorescence imaging enable classification of bone perfusion in patients undergoing leg amputation.

Dynamic contrast-enhanced fluorescence imaging (DCE-FI) classification of tissue viability in twelve adult patients undergoing below knee leg amputation is presented. During amputation and with the distal bone exposed, indocyanine green contrast-enhanced images were acquired sequentially during baseline, following transverse osteotomy and following periosteal stripping, offering a uniquely well-controlled fluorescence dataset. An unsupervised classification machine leveraging 21 different spatiotemporal features was trained and evaluated by cross-validation in 3.5 million regions-of-interest obtained from 9 patients, demonstrating accurate stratification into normal, suspicious, and compromised regions. The machine learning (ML) approach also outperformed the standard method of using fluorescence intensity only to evaluate tissue perfusion by a two-fold increase in accuracy. The generalizability of the machine was evaluated in image series acquired in an additional three patients, confirming the stability of the model and ability to sort future patient image-sets into viability categories.