Korean Society of Endourology and Robotics (KSER) recommendation on the diagnosis, treatment, and prevention of urolithiasis.

This article provides evidence-based recommendations and expert opinions to aid urologists in making optimal decisions regarding managing urolithiasis in various clinical scenarios. The most frequently asked questions by urologists in their clinical practice have been collected and answered in the form of FAQs; based on the latest evidence and expert opinions. The natural history of urolithiasis is divided into active treatment and silent phases, with the active treatment stage divided into typical and special situations and peri-treatment management. The authors address 28 key questions, offering practical guidance for the proper diagnosis, treatment, and prevention of urolithiasis in clinical practice. This article is expected to be served as a valuable resource for urologists.

Fasciclin 2 plays multiple roles in promoting cell migration within the developing nervous system of Manduca sexta.

The coordination of neuronal and glial migration is essential to the formation of most nervous systems, requiring a complex interplay of cell-intrinsic responses and intercellular guidance cues. During the development of the enteric nervous system (ENS) in Manduca sexta (tobacco hornworm), the IgCAM Fasciclin 2 (Fas2) serves several distinct functions to regulate these processes. As the ENS forms, a population of 300 neurons (EP cells) undergoes sequential phases of migration along well-defined muscle pathways on the visceral mesoderm to form a branching Enteric Plexus, closely followed by a trailing wave of proliferating glial cells that enwrap the neurons. Initially, both the neurons and glial cells express a GPI-linked form of Fas2 (GPI-Fas2), which helps maintain cell-cell contact among the pre-migratory neurons and later promotes glial ensheathment. The neurons then switch isoforms, predominantly expressing a combination of transmembrane isoforms lacking an intracellular PEST domain (TM-Fas2 PEST-), while their muscle band pathways on the midgut transiently express transmembrane isoforms containing this domain (TM-Fas2 PEST+). Using intracellular injection protocols to manipulate Fas2 expression in cultured embryos, we found that TM-Fas2 promotes the directed migration and outgrowth of individual neurons in the developing ENS. Concurrently, TM-Fas2 expression by the underlying muscle bands is also required as a substrate cue to support normal migration, while glial expression of GPI-Fas2 helps support their ensheathment of the migratory neurons. These results demonstrate how a specific IgCAM can play multiple roles that help coordinate neuronal and glial migration in the developing nervous system.

[A Case of Epiploic Appendagitis after COVID-19].

Acute epiploic appendagitis is an uncommon cause of abdominal pain resulting from appendageal ischemia caused by torsion or thrombosis of the draining vein. It is frequently misdiagnosed as acute appendicitis or diverticulitis. The coronavirus disease 2019 (COVID-19) pandemic has changed how this rare disease is diagnosed. There was a report of a young men diagnosed with COVID-19 and epiploic appendagitis as a rare cause of abdominal pain. In addition, a 50-year-old men was diagnosed with epiploic appendagitis during the treatment of COVID-19. This paper reports the case of a 53-year-old men who presented with right lower quadrant abdominal pain after COVID-19 and was diagnosed with acute epiploic appendagitis by computed tomography image findings. The thrombotic condition of COVID-19 may contribute to acute appendagitis, but more studies are needed to confirm this hypothesis.

Commentary on "Novel Interaction of the Dopamine D2 Receptor and the Ca2+ Binding Protein S100B: Role in D2 Receptor Function".

We previously proposed that the dopamine D2 receptor-interacting protein S100B binds to a putative S100B-binding motif at residues R233-L240 toward the N terminus of the third intracellular loop. We used in vitro pull-down assays with FLAG-tagged fragments of the rat dopamine D2 receptor third intracellular loop (D2-IC3) and in vitro-synthesized S100B to evaluate this hypothesis. Our results indicate that the putative S100B-binding motif is neither necessary nor sufficient for strong binding of S100B to D2-IC3. Instead, two residues at the junction of the fifth membrane-spanning domain and the cytoplasmic extension of that α-helical domain, K211-I212, are required for robust, calcium-sensitive binding of S100B. This is also the approximate location of previously identified determinants for the binding of arrestin and calmodulin. A D2 receptor mutation converting I212 to phenylalanine has been described in patients with a hyperkinetic movement disorder. SIGNIFICANCE STATEMENT: S100B is a small calcium-binding protein that modulates signaling by the dopamine D2 receptor. New data suggest that the previous hypothesis about the involvement of an S100B-binding motif is incorrect, and that an important determinant of S100B binding includes a residue that is mutated in patients with a hyperkinetic movement disorder.

How to Improve Projection in Nipple Reconstruction: A Modified Method Using Acellular Dermal Matrix Disk and Fragments.

BACKGROUND: Nipple reconstruction is an essential, final stage in breast reconstruction. However, postoperative reduction in nipple projection often results in low patient satisfaction. The authors studied the causes of the projection decline and developed a new method using acellular dermal matrix. This research studies the effectiveness of the new method. METHODS: The nipple flap was elevated using a modified C-V flap, and acellular dermal matrix disk was fixed onto the floor. A column was made, into which acellular dermal matrix fragments were put in to retain the projection. The footprint diameter and projection at 1 year were compared with those of the control group, in which acellular dermal matrix was not used. The authors studied the correlation between diameter and projection and whether reconstruction method caused any impact. RESULTS: At 1-year follow-up, the nipple diameter and projection in the acellular dermal matrix group were measured to be 102.90 percent and 64.19 percent, respectively, of the baseline. Compared with the control group, the diameter was significantly smaller (p = 0.00) and the projection was higher (p = 0.00). A significant correlation was identified between nipple diameters and projections, at 1-year follow-up, across the total 90 reconstructed nipples (p = 0.00). Different reconstruction methods did not show significant differences in terms of nipple diameter and projection, but the projections at 1 year were highest in the latissimus dorsi flap plus implant group, followed by the expander group and the transverse rectus abdominis musculocutaneous flap group. CONCLUSION: Nipple reconstruction using acellular dermal matrix disk and fragments prevents downward shifting of the nipple tissue and broadening of the footprint diameter and thus is favorable for long-term maintenance of nipple projection.

An Ideal Female Breast Shape in Balance with the Body Proportions of Asians.

Defining an ideal breast shape is one of the most fundamental and essential parts for a breast surgery. To propose a set of criteria for determining an ideal breast shape of Asians, the authors performed a survey using a questionnaire based on important esthetic elements of a breast. METHODS: The authors created a 11-item questionnaire, asking breast shape preference in the frontal and the lateral views. Each question had multiple options, each of which was accompanied by adequate illustrations. RESULTS: A total of 1,012 Asian responses were collected. In the frontal view, preferences converged for position of the nipple to be at 45% of the SU (distance from the sternal notch to the umbilicus) and the inframammary fold at 60% of the SU. For lateral bulging of the breast, the respondents preferred it to be 100% of the upper buttock, and 100% of the interacromion width. As for the lower pole height, breast width ratio of 50% was the most preferred. In the lateral view, straight slope of the upper breast was the most preferred, along with a 1.0 projection ratio and a front-facing nipple. The most ideal vertical proportion of the breast footprint was selected as 65:35, and for the anterior breast as 55:45. CONCLUSION: The authors used a questionnaire analysis, which considers a proportional balance between the breast and the whole body shape, and proposed that an ideal breast shape can be used effectively in planning for and assessing the outcomes of breast surgery.

Selective isolation and noninvasive analysis of circulating cancer stem cells through Raman imaging.

Circulating cancer stem cells (CCSCs), a rare circulating tumor cell (CTC) type, recently arose as a useful resource for monitoring and characterizing both cancers and their metastatic derivatives. However, due to the scarcity of CCSCs among hematologic cells in the blood and the complexity of the phenotype confirmation process, CCSC research can be extremely challenging. Hence, we report a nanoparticle-mediated Raman imaging method for CCSC characterization which profiles CCSCs based on their surface marker expression phenotypes. We have developed an integrated combinatorial Raman-Active Nanoprobe (RAN) system combined with a microfluidic chip to successfully process complete blood samples. CCSCs and CTCs were detected (90% efficiency) and classified in accordance with their respective surface marker expression via completely distinct Raman signals of RANs. Selectively isolated CCSCs (93% accuracy) were employed for both in vitro and in vivo tumor phenotyping to identify the tumorigenicity of the CCSCs. We utilized our new method to predict metastasis by screening blood samples from xenograft models, showing that upon CCSC detection, all subjects exhibited liver metastasis. Having highly efficient detection and noninvasive isolation capabilities, we have demonstrated that our RAN-based Raman imaging method will be valuable for predicting cancer metastasis and relapse via CCSC detection. Moreover, the exclusion of peak overlapping in CCSC analysis with our Raman imaging method will allow to expand the RAN families for various cancer types, therefore, increasing therapeutic efficacy by providing detailed molecular features of tumor subtypes.

The Measurement of Palpebral Fissure Height Using the Intersection Angle (the Réal Angle) Between the Meeting Points of the Upper Eyelid and the Edge of the Cornea.

BACKGROUND: We evaluated a new palpebral fissure height measurement to evaluate medial, lateral, and overall ptosis. METHODS: We photographed 250 Koreans (44 males, 206 females) and evaluated their Réal 1 angle (angle between the meeting points of the upper eyelid and the corneal edge), Réal 2 angle (angle between the meeting point of the upper eyelid, medial corneal edge and a vertical line through the center of the pupil), Réal 3 angle (angle between the meeting point of the upper eyelid, lateral corneal edge and a vertical line through the center of the pupil), and Réal 4 angle (Réal 2-Réal 3). Angles were compared between sexes and age groups. We then evaluated the Réal angles of 13 Korean actresses. RESULTS: Mean age was 31.85 ± 14.60 years; Réal 1 was 129.01° ± 14.23°, Réal 2 was 68.20° ± 7.49°, Réal 3 was 60.80° ± 9.65°. There was no significant difference between the sexes in Réal 1, Réal 2, and Réal 3 angles. Réal 1 increased with age, and Réal 4 decreased with age. All Réal angles were significantly different between age groups. The actresses' mean age was 30.66 ± 8.01 years; Réal 1 was 102.84° ± 10.16°, Réal 2 was 57.87° ± 6.10°, and Réal 3 was 44.97° ± 8.74°. CONCLUSION: This simple measurement of palpebral fissure height using Réal angles consistently evaluated the amount of medial, lateral, and general ptosis. For average Korean eyes, the lateral portion of the upper eyelid is slightly higher than the medial portion; however, this lateral portion droops with age. Korean actresses have vertically higher eyes than average Korean women. LEVEL OF EVIDENCE V: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

ΔN-ASPP2, a novel isoform of the ASPP2 tumor suppressor, promotes cellular survival.

ASPP2 is a tumor suppressor that works, at least in part, through enhancing p53-dependent apoptosis. We now describe a new ASPP2 isoform, ΔN-ASPP2, generated from an internal transcription start site that encodes an N-terminally truncated protein missing a predicted 254 amino acids. ΔN-ASPP2 suppresses p53 target gene transactivation, promoter occupancy, and endogenous p53 target gene expression in response to DNA damage. Moreover, ΔN-ASPP2 promotes progression through the cell cycle, as well as resistance to genotoxic stress-induced growth inhibition and apoptosis. Additionally, we found that ΔN-ASPP2 expression is increased in human breast tumors as compared to adjacent normal breast tissue; in contrast, ASPP2 is suppressed in the majority of these breast tumors. Together, our results provide insight into how this new ASPP2 isoform may play a role in regulating the ASPP2-p53 axis.

Development of visual peak selection system based on multi-ISs normalization algorithm to apply to methamphetamine impurity profiling.

The aim of this study is to improve resolution of impurity peaks using a newly devised normalization algorithm for multi-internal standards (ISs) and to describe a visual peak selection system (VPSS) for efficient support of impurity profiling. Drug trafficking routes, location of manufacture, or synthetic route can be identified from impurities in seized drugs. In the analysis of impurities, different chromatogram profiles are obtained from gas chromatography and used to examine similarities between drug samples. The data processing method using relative retention time (RRT) calculated by a single internal standard is not preferred when many internal standards are used and many chromatographic peaks present because of the risk of overlapping between peaks and difficulty in classifying impurities. In this study, impurities in methamphetamine (MA) were extracted by liquid-liquid extraction (LLE) method using ethylacetate containing 4 internal standards and analyzed by gas chromatography-flame ionization detection (GC-FID). The newly developed VPSS consists of an input module, a conversion module, and a detection module. The input module imports chromatograms collected from GC and performs preprocessing, which is converted with a normalization algorithm in the conversion module, and finally the detection module detects the impurities in MA samples using a visualized zoning user interface. The normalization algorithm in the conversion module was used to convert the raw data from GC-FID. The VPSS with the built-in normalization algorithm can effectively detect different impurities in samples even in complex matrices and has high resolution keeping the time sequence of chromatographic peaks the same as that of the RRT method. The system can widen a full range of chromatograms so that the peaks of impurities were better aligned for easy separation and classification. The resolution, accuracy, and speed of impurity profiling showed remarkable improvement.

Estimation of the synthetic routes of seized methamphetamines using GC-MS and multivariate analysis.

One hundred and twenty six seized methamphetamine (MA) samples were analyzed using GC-MS. All the peaks that appeared in the chromatograms were investigated and 61 impurities including n-octacosane (internal standard) were identified. Among them, 37 impurities were already known or newly identified by comparing with commercial library entries and 18 impurities were detected for the first time. To estimate the synthetic routes of MA samples, route specific impurities had to be selected for each method. Two naphthalenes, 1,3-dimethyl-2-phenylnaphthalene and 1-benzyl-3-methylnaphthalene were selected as Nagai route specific impurities and three diasteromers, UK-19.62(58_165_178) I, UK-19.95(58_165_178) II, UK-20.49(58_165_178) III were also selected not only for their high frequency detection only in Nagai samples but also for the high principal component analysis (PCA) correlation values. For the Emde route, N,N-dimethyl-3,4-diphenylhexane-2,5-diamine and N-methyl-1-{4-[2-(methylamino)propyl]phenyl}-1-phenylpropan-2-amine were selected as route specific impurities, and N,N-di(ß-phenylisopropyl)amine I (DPIA I), N,N-di(ß-phenylisopropyl)amine II (DPIA II), N,N-di(ß-phenylisopropyl)methylamine I (DPIMA I) and N,N-di(ß-phenylisopropyl)methylamine II (DPIMA II) were selected for the Leuckart route. With these route specific impurities, synthetic routes could be identified for 78 of the 126 samples. The 61 impurities were registered in AMDIS target component library and the GC-MS data were deconvoluted. After AMDIS deconvolution, a matrix file was composed and then multivariate analyses were performed to estimate the synthetic route for unknown samples. The unsupervised methods, hierarchical clustering analysis (HCA) and PCA clustered the samples according to the closeness between samples. Two classification functions were obtained from discriminant analysis (DA) and the synthetic routes of the unknown samples were predicted using these two functions.

Simultaneous capture and in situ analysis of circulating tumor cells using multiple hybrid nanoparticles.

Using hybrid nanoparticles (HNPs), we demonstrate simultaneous capture, in situ protein expression analysis, and cellular phenotype identification of circulating tumor cells (CTCs). Each HNP consists of three parts: (i) antibodies that bind specifically to a known biomarker for CTCs, (ii) a quantum dot that emits fluorescence signals, and (iii) biotinylated DNA that allows capture and release of CTC-HNP complex to an in-house developed capture & recovery chip (CRC). To evaluate our approach, cells representative of different breast cancer subtypes (MCF-7: luminal; SK-BR-3: HER2; and MDA-MB-231: basal-like) were captured onto CRC and expressions of EpCAM, HER2, and EGFR were detected concurrently. The average capture efficiency of CTCs was 87.5% with identification accuracy of 92.4%. Subsequently, by cleaving the DNA portion with restriction enzymes, captured cells were released at efficiencies of 86.1%. Further studies showed that these recovered cells are viable and can proliferate in vitro. Using HNPs, it is possible to count, analyze in situ protein expression, and culture CTCs, all from the same set of cells, enabling a wide range of molecular- and cellular-based studies using CTCs.

A trachea-inspired bifurcated microfilter capturing viable circulating tumor cells via altered biophysical properties as measured by atomic force microscopy.

Circulating tumor cells (CTCs), though exceedingly rare in the blood, are nonetheless becoming increasingly important in cancer diagnostics. Despite this keen interest and the growing number of potential clinical applications, there has been limited success in developing a CTC isolation platform that simultaneously optimizes recovery rates, purity, and cell compatibility. Herein, a novel tracheal carina-inspired bifurcated (TRAB) microfilter system is reported, which uses an optimal filter gap size satisfying both 100% theoretical recovery rate and purity, as determined by biomechanical analysis and fluid-structure interaction (FSI) simulations. Biomechanical properties are also used to clearly discriminate between cancer cells and leukocytes, whereby cancer cells are selectively bound to melamine microbeads, which increase the size and stiffness of these cells. Nanoindentation experiments are conducted to measure the stiffness of leukocytes as compared to the microbead-conjugated cancer cells, with these parameters then being used in FSI analyses to optimize the filter gap size. The simulation results show that given a flow rate of 100 µL min(-1), an 8 µm filter gap optimizes the recovery rate and purity. MCF-7 breast cancer cells with solid microbeads are spiked into 3 mL of whole blood and, by using this flow rate along with the optimized microfilter dimensions, the cell mixture passes through the TRAB filter, which achieves a recovery rate of 93% and purity of 59%. Regarding cell compatibility, it is verified that the isolation procedure does not adversely affect cell viability, thus also confirming that the re-collected cancer cells can be cultured for up to 8 days. This work demonstrates a CTC isolation technology platform that optimizes high recovery rates and cell purity while also providing a framework for functional cell studies, potentially enabling even more sensitive and specific cancer diagnostics.

Highly efficient assay of circulating tumor cells by selective sedimentation with a density gradient medium and microfiltration from whole blood.

Isolation of circulating tumor cells (CTCs) by size exclusion can yield poor purity and low recovery rates, due to large variations in size of CTCs, which may overlap with leukocytes and render size-based filtration methods unreliable. This report presents a very sensitive, selective, fast, and novel method for isolation and detection of CTCs. Our assay platform consists of three steps: (i) capturing CTCs with anti-EpCAM conjugated microbeads, (ii) removal of unwanted hematologic cells (e.g., leukocytes, erythrocytes, etc.) by selective sedimentation of CTCs within a density gradient medium, and (iii) simple microfiltration to collect these cells. To demonstrate the efficacy of this assay, MCF-7 breast cancer cells (average diameter, 24 µm) and DMS-79 small cell lung cancer cells (average diameter, 10 µm) were used to model CTCs. We investigated the relative sedimentation rates for various cells and/or particles, such as CTCs conjugated with different types of microbeads, leukocytes, and erythrocytes, in order to maximize differences in the physical properties. We observed that greater than 99% of leukocytes in whole blood were effectively removed at an optimal centrifugal force, due to differences in their sedimentation rates, yielding a much purer sample compared to other filter-based methods. We also investigated not only the effect of filtration conditions on recovery rates and sample purity but also the sensitivity of our assay platform. Our results showed a near perfect recovery rate (~99%) for MCF-7 cells and very high recovery rate (~89%) for DMS-79 cells, with minimal amounts of leukocytes present.

Sensitive and simultaneous detection of cardiac markers in human serum using surface acoustic wave immunosensor.

We present a rapid and sensitive surface acoustic wave (SAW) immunosensor that utilizes gold staining as a signal enhancement method. A sandwich immunoassay was performed on sensing area of the SAW sensor, which could specifically capture and detect cardiac markers (cardiac troponin I (cTnI), creatine kinase (CK)-MB, and myoglobin). The analytes in human serum were captured on gold nanoparticles (AuNPs) that were conjugated in advance with detection antibodies. Introduction of these complexes to the capture antibody-immobilized sensor surface resulted in a classic AuNP-based sandwich immunoassay format that has been used for signal amplification. In order to achieve further signal enhancement, a gold staining method was performed, which demonstrated that it is possible to obtain gold staining-mediated signal augmentation on a mass-sensitive device. The sensor response due to gold staining varied as a function of cardiac marker concentration. We also investigated effects of increasing operating frequency on sensor responses. Results showed that detection limit of the SAW sensor could be further improved by increasing the operating frequency.

Clinical characteristics and surgical outcomes in adolescents and adults with varicocele.

PURPOSE: To compare clinical characteristics and surgical results in adolescents and adults with varicocele. MATERIALS AND METHODS: We retrospectively analyzed the characteristics of 93 patients, 34 adolescents (mean age, 14.4±2.1 years) and 59 adults (mean age, 30.4±12.4 years), who underwent surgical repair of varicocele between 2006 and 2009. Median follow-up time in all patients was 18.7 months. The most bothersome symptoms, bilaterality, grades, surgical methods, artery-sparing rates, operation times, semen analysis, success rates, and recurrence-free period were compared between the two groups. RESULTS: The overall success rate of surgical repair was 92.5%. The most bothersome symptoms were scrotal mass, pain, and hypotrophy in adolescents and pain, scrotal mass, infertility, and hypotrophy in adults (p=0.008). There were no significant between-group differences in bilaterality, grades, surgical methods, operation times, pre- or postoperative semen analyses, success rates, or recurrence-free periods. Patients who underwent artery-sparing surgery had higher recurrence rates than did those who underwent surgery that did not spare arteries. In adults, semen density increased significantly after surgery, from 35.6 million/ml to 49.6 million/ml (p=0.046). CONCLUSIONS: There were no significant differences in clinical characteristics or surgical results between adolescents and adults with varicocele, except for the most bothersome symptoms. Semen density increased after surgery in both groups.

Electrochemical cell lysis device for DNA extraction.

We present a novel electrochemical cell lysis device to prepare DNA samples for lab-on-a-chip (LOC) applications. It utilizes the electrolysis of saline solution to generate hydroxide ions (OH(-)) at the cathode as alkaline lytic agents. Cathode and anode chambers are separated by a negatively-charged ion exchangeable polymer diaphragm to maintain the high pH level for efficient cell lysis in the cathode chamber, to prevent inflow of PCR-amplification inhibitors from the anode chamber, and to minimize binding of DNA molecules. Electric current flow and pH maintenance, which depended on the device design, were two important parameters of the device performance. After optimizing the design and visually confirming cell lysis of Chinese hamster ovary (CHO) cells in a very short amount of time, we directly electrolyzed four bacterial cell types suspended in saline solution. Real-time PCR (qPCR) analysis showed that our device could lyse both gram-positive and gram-negative bacterial cells with higher efficiency than other common methods and could detect DNA on the microlitre scale. Our data demonstrate several advantages of the proposed device: absence of cell lysis chemicals and heating; no adverse effects on PCR amplification; low DNA loss; low voltage and power consumption; and rapid processing. The device could potentially be applied as an on-chip DNA extraction component.

Surface acoustic wave immunosensor for real-time detection of hepatitis B surface antibodies in whole blood samples.

We demonstrate an application of Love wave mode surface acoustic wave (SAW) immunosensor to detect hepatitis B surface antibody (HBsAb) in aqueous conditions. SiO(2) guiding layer was deposited on 36 degrees YX-LiTaO(3) piezoelectric single crystal substrate to protect the electrodes and to trap the acoustic energy near the surface, and hepatitis B surface antigen (HBsAg) was immobilized on the sensing area. The resonance frequency shift was monitored to detect specific binding of HBsAb to immobilized HBsAg. To eliminate the effects of other physical factors except for the mass change, the resonance frequency was compared to that of a reference SAW device coated with bovine serum albumin (BSA) to block binding of HBsAb. The guiding layer thickness with maximum mass sensitivity was found to be 5 microm, which was in agreement with the theoretical calculation, and the center resonance frequency was around 199 MHz. The sensor showed binding specificity to HBsAb and a linear relationship between the frequency shift and the antibody concentration with sensitivity of 0.74 Hz/(pg/microl) and detection limit below 10 pg/microl. In addition, our SAW immunosensor successfully detected HBsAb in whole blood samples without any pretreatment, opening up its applicability in fast label-free protein detection methods.

Application of complement 1q for the site-selective recognition of immune complex in protein chip.

Complement 1q (C1q) was applied for the specific recognition of antibody-antigen complex in antibody-based protein chip. The specific binding of C1q to antibody-antigen complex was investigated by surface plasmon resonance (SPR) with respect to Yersinia entericolitica, Salmonella typimurium, insulin, and bovine serum albumin. The protein chip was fabricated with two different kinds of antibodies a zigzag configuration. When one of antigens and fluorescein-isothiocyanate (FITC)-labeled C1q was applied on the protein chip, the specific binding event of C1q to immune complexes formed on protein chip was observed by fluorescence microscopy. These results implicate that the C1q can be used as an alternative to many antibodies that may be utilized individually on each spot of the protein chip.