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Authors: Sean Sterrenberg, David Gallacher, Melissa Schmidt, Rebecca Rugel, Summer Gerke, Kenneth Gundle, Ryan Wallenberg
Group: Smokerlyzer Study, PVAMC
Summary: Background: Tobacco use is associated with increased post-operative complications and is often underreported by patients. Traditional biochemical testing methods, including serum and urine cotinine assays, do not differentiate active smoking from nicotine replacement therapy (NRT). This study intends to evaluate the effectiveness of a novel point-of-care carbon monoxide (CO) breath test to verify smoking status and differentiate active smoking from NRT pre-operatively in a cohort of veteran orthopaedic patients. Methods: This is a single-institution, prospective cohort study of adult orthopaedic patients at the Portland VA Medical Center to implement and evaluate a point-of-care CO breath test, the Smokerlyzer® Micro EC50, for pre-operative smoking status verification. Patients who are indicated for orthopaedic surgery in the pre-operative clinic will be offered inclusion. Self-reported smoking status will be obtained pre-operatively to categorize patients as non-smoker, quitters, or active smokers. Exhaled CO and serum cotinine levels will be obtained at the initial clinic visit, pre-operative visit, and day of surgery. Primary endpoints are concordance between self-reported smoking status and CO levels, and concordance between exhaled CO and serum cotinine levels.Discussion: Pre-operative smoking cessation interventions have been shown to decrease smoking rates on the day of surgery and promote abstinence 30 days post-operatively. NRTs are commonly used to promote smoking cessation, however their effect on surgical outcomes are not known. This study aims to validate the use of point-of-care testing that differentiates active smoking from NRT, and identify patients at risk for smoking status misclassification. Results would provide preliminary data to directly test NRT use, separate from tobacco use, and surgical outcomes.Specific Aims: 1. To determine if expired air CO levels can be used to identify patients that are at risk of smoking status misclassification pre-operatively, including patients that are on nicotine replacement therapy. 2. To evaluate the concordance between self-reported smoking and expired air CO levels in the veteran orthopedic population. 3. To evaluate the concordance between pre-operative expired air CO and typical serum levels in the orthopedic population. Hypotheses: 1. Expired air CO testing will allow for differentiation between active smokers and those on nicotine replacement therapy compared to serum cotinine testing. 2. Self-reported “quitters” will have lower agreement between self-reported smoking status and biological testing compared to self-reported “non-smokers” and “active smokers.”3. Compared to serum cotinine testing, expired air CO testing will allow for non-inferior accuracy in verifying smoking status.

Proper citation: Sean Sterrenberg, David Gallacher, Melissa Schmidt, Rebecca Rugel, Summer Gerke, Kenneth Gundle, Ryan Wallenberg 2019. Is Expired Air Carbon Monoxide Testing Effective for Screening of Cigarette Use in Orthopaedic Patients?. protocols.io https://dx.doi.org/10.17504/protocols.io.7crhiv6 Copy   

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Authors: Froylan Sosa, Peter Hansen
Summary: This is a simple protocol for collecting skin biopsies from cattle. The procedure described here was used to collect skin for histological analysis but it could also be used to collect skin for PCR or other purposes. Our experience indicates that there are few complications with the procedure. The most common is that occasionally there is a slight amount of bleeding that can be treated by applying pressure using a gauze. Healing occurs rapidly and biopsy sites are difficult to detect 14 days after the procedure. The protocol is best read by using the attached pdf file.

Proper citation: Froylan Sosa, Peter Hansen 2020. Collection of skin biopsies from cattle. protocols.io https://dx.doi.org/10.17504/protocols.io.bm3yk8pw Copy   

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Authors: Anna Racioppi, Tamara Sheinbaum, Georgina M. Gross, Sergi Ballespí, Thomas R. Kwapil, Neus Barrantes-Vidal
Summary: The present study forms part of an ongoing longitudinal project examining risks and resilience for schizophrenia-spectrum disorders in a non-clinical ascertained sample of college students. The main objective of the study was examine the validity of psychometrically assessed positive and negative schizotypy at a three-year follow-up assessment (Time 3; T3) of the sample initially reported in the cross-sectional study of Barrantes-Vidal et al. (2013). Participants were recruited from the undergraduate population enrolled in Psychology courses at the Universitat Autònoma de Barcelona (UAB). A group of 589 unselected students completed a mass screening assessment of self-reported measures at Time 1 (T1). Participants were administered the Wisconsin Schizotypy Scales (WSS) intermixed with an infrequency scale, the positive symptoms subscale of the Community Assessment of Psychic Experiences (CAPE), and the suspiciousness subscale of the Schizotypal Personality Questionnaire (SPQ). Usable screening data were obtained from 547 students (mean age = 20.6; SD = 4.1; 86% female, 42 were excluded from the final study due to invalid protocols). Two interview and self-report measures assessments were conducted after T1. The Time 2 (T2) and Time 3 (T3) interviews were made by psychologists and advanced graduate students in clinical psychology. All interviewers were extensively trained and were unaware of participants' scores on the T1 and T2 measures. Of the 547 students who completed T1 assessment, a subset of 339 was invited to participate at T2 assessment with the aim of assessing 200 individuals. In order to ensure adequate inclusion of participants reporting high levels of schizotypy, the students group included 189 who had elevated scores (standard scores based upon sample norms of at least 1.0) on the positive or negative schizotypy factors derived from the WSS, or the SPQ-suspiciousness, or the CAPE, and 150 randomly selected participants who had standard scores below 1.0 on each of these measures.A total of 214 students (mean age = 21.4 years; SD = 2.4; 78% female) completed T2 assessment which took place an average of 1.7 years (SD= 0.2 years, range 1.4 to 2.2 years) after T1. This sample included 123 participants with at least one schizotypy screening score above 1.0 and 91 with standard scores below 1.0. The T3 assessment took place an average of 1.4 years (SD= 0.3 years, range 0.9 to 2.1 years) after T2, and of 3.1 years (SD= 0.3 years, range 2.6 to 3.6 years) after T1. At T3, due to financial limitation we targeted a subset of 134 students (93 with high schizotypy and 41 with standard scores below 1.0). Of these, 103 (77%) participants (mean age= 23.06; SD=2.6; 37.9% male) were interviewed at T3. The sample included 75 of 93 (82%) participants with elevated schizotypy scores and 28 of 43 (65%) with standard scores below 1.0. There were no significant differences on positive or negative schizotypy scores between the participants assessed at T3 and the non-followed participants. This study was conducted according to guidelines of the University Ethics Committee of the UAB and participants provided informed consent at each assessment.

Proper citation: Anna Racioppi, Tamara Sheinbaum, Georgina M. Gross, Sergi Ballespí, Thomas R. Kwapil, Neus Barrantes-Vidal 2018. Protocol of the Psychometric High-Risk Strategy Project for Examining Risk and Resilience Trajectories across the Psychosis Continuum. protocols.io https://dx.doi.org/10.17504/protocols.io.ubieske Copy   

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Authors: Wyatt Million, Carly Kenkel
Group: Coral Bleaching RCN protocols
Summary: This is a protocol for generating images to be used in 3D model building via Agisoft Metashape for coral photogrametry. This will cover underwater, field-based methods and tips to collect photographs and pre-processing of photos to improve model building. Image capture is the most important part of 3D photogrammetry because the photos taken at this point will be all that you'll have to build models and collect data. As such, you want to ensure you have enough photos to work with in the future so, in general, more is better. That being said, too many blurry or out of focus pictures will hamper model building. You can optimize your time in the field by taking enough photos from the appropriate angles, however efficiency will come with practice. This is the protocol developed and used by the Kenkel lab to phenotype Acropora cervicornis colonies as part of field operations in the Florida Keys. We incorporate Agisoft Metashape markers in this workflow to scale models and improved model building. The scaling objects used by the Kenkel lab are custom-made, adjustable PVC arrays that include unique markers and bleaching color cards, affectionately called the "Tomahawk". Specs for building a Tomahawk are included in this protocol. Filtering and pre-processing of photos is not always necessary but can be used to salvage 3D models that would be otherwise blurry or incomplete. Here, we describe photo editing in Adobe Lightroom to adjust several characteristics of hundreds of images simultaneously.For a walkthrough and scripts to run Agisoft Metashape on the command line, see https://github.com/wyattmillion/Coral3DPhotogram. For directions to phenotype coral from 3D models see our Phenotyping in MeshLab protocol.These protocols, while created for branching coral, can be applied to 3D models of any coral morphology or any object really. Our goal is to make easy-to-use protocols using accessible softwares in the hopes of creating a standardized method for 3D photogrammetry in coral biology.

Proper citation: Wyatt Million, Carly Kenkel 2020. Image capture and pre-filtering for 3D photogrammetry of coral colonies. protocols.io https://dx.doi.org/10.17504/protocols.io.bgdcjs2w Copy   

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Authors: Joshua Welsh, Jennifer Jones
Group: Translational Nanobiology Section
Summary: This protocol outlines the steps required to catalogue fluorescence reference materials using the FCMPASS software. This is one of a number of protocols in the pipeline for performing small particle calibration using the fcmpass software package.

Proper citation: Joshua Welsh, Jennifer Jones 2020. FCMPASS - Cataloguing fluorescence reference materials. protocols.io https://dx.doi.org/10.17504/protocols.io.bhvvj666 Copy   

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Authors: Sarah Piccirillo, Melissa G. White, Jeffrey C. Murphy, Douglas J. Law, and Saul M. Honigberg
Group: Genetics
Summary: The following is a modification of a Scherz R., 2001 method for colony embedding. The protocol described here is from:Sarah Piccirillo, et. al. The Rim101p/PacC Pathway and Alkaline pH Regulate Pattern Formation in Yeast Colonies(2010)Genetics184:707-716; doi:10.1534/genetics.109.113480Please see the full manuscript for additional details.

Proper citation: Sarah Piccirillo, Melissa G. White, Jeffrey C. Murphy, Douglas J. Law, and Saul M. Honigberg 2015. Embedding yeast colonies for light and electron microscopy. protocols.io https://dx.doi.org/10.17504/protocols.io.dw97h5 Copy   

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Authors: 张 雪
Group: Zhang Xue

Proper citation: 张 雪 2020. total RNA抽提. protocols.io https:// Copy   

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Authors: Chu-Yu Chin, Sun-Yuan Hsieh, Vincent S. Tseng
Summary: The early assessment of disease risk is an emerging topic in medical informatics. If diseases are detected at an early stage, prognosis can be improved and medical resources can be used more efficiently. A number of recent studies have considered risk factor analysis approaches, such as association rule mining, sequential rule mining, regression, and medical expert advice. In this study, for improving disease risk assessment, non-negative matrix factorization and support vector machine (SVM) were integrated to discover important and implicit risk factors.To make the method easy to follow, here we provide an experimental protocal. This experimental protocal comprises three main stages: data preprocessing, risk factor optimization, and early disease risk assessment. To discover the optimized risk factors, the NMF algorithm with parameter optimization was used for constructing the NMF-based matrix. In the assessment model learning and early disease risk assessment stages, the machine learning classifier SVM was used for disease modeling with the NMF-based matrix, yielding the final disease risk assessment, which serves as an excellent reference for physicians and patients.

Proper citation: Chu-Yu Chin, Sun-Yuan Hsieh, Vincent S. Tseng 2018. Effective early disease risk assessment with matrix factorization on a large-scale medical database. protocols.io https://dx.doi.org/10.17504/protocols.io.rv2d68e Copy   

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Authors: James Lloyd, Ryan Lister
Summary: To extract plasmid DNA from Agrobacterium, to confirm the correct plasmid has been taken-up, this protocol allows you to directly perform a mini-prep on transformed Agrobacterium, digest this DNA and visualize it on a gel. This is without the need of rescuing the plasmid in E. coli or genotyping with PCR. This protocol using the commercially available Bioline ISOLATE II Plasmid Mini Kit, designed for use with E. coli. The standard protocol this is modified from is available here: https://www.bioline.com/au/downloads/dl/file/id/1219/isolate_ii_plasmid_mini_kit_protocol.pdf

Proper citation: James Lloyd, Ryan Lister 2018. Extracting plasmid DNA from Agrobacterium with the Bioline ISOLATE II Plasmid Mini Kit. protocols.io https://dx.doi.org/10.17504/protocols.io.s8fehtn Copy   

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Authors: Lenny Teytelman
Group: VERVE Net
Summary: This is a highly reproducible method for creating hit science videos. Guaranteed to lead to millions of views, fame, faculty position, tenure, and stable funding.* Legal disclaimer: This is not a peer reviewed protocol. Therefore, all claims should be taken with as much skepticism as anything that is peer reviewed.

Proper citation: Lenny Teytelman 2016. Making a hit science video. protocols.io https://dx.doi.org/10.17504/protocols.io.emqbc5w Copy   

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Authors: Sebastian Triesch, Maximilian Dietsch
Group: Axmann Lab, CyanoWorld, M4455 - Synthetische Biologie und Biotechnologie
Summary: The following protocol is optimized to generate first-strand cDNA for use in two step-PCR.

Proper citation: Sebastian Triesch, Maximilian Dietsch 2019. First strand cDNA synthesis (ThermoScientific RevertAid). protocols.io https://dx.doi.org/10.17504/protocols.io.ze3f3gn Copy   

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Authors: Kelsey Miller, Sam Li
Group: BioLegend
Summary: BioLegend MojoSort™ nanobeads work in commonly used separation columns, based on our internal research as well as validation by external testing by academic labs. This simple protocol consists of following the MojoSort™ protocol to label the cells with pre-diluted MojoSort™ reagents and using the columns as indicated by the manufacturer.Note: Due to the properties of our beads, it may be possible to use far fewer beads and less antibody than with other commercial suppliers. We recommend a titration to find the best dilution factor. However, as a general rule, dilutions ranging from 1:3 to 1:20 for the Nanobeads can be used. Please contact BioLegend Technical Service ([email protected]) if further assistance is needed.

Proper citation: Kelsey Miller, Sam Li 2019. MojoSort™ Streptavidin Nanobeads Column Protocol - Positive Selection. protocols.io https://dx.doi.org/10.17504/protocols.io.692hh8e Copy   

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Authors: USDA
Group: PDI Test

Proper citation: USDA 2021. General Lab SOP. protocols.io https:// Copy   

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Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: This SOP provides instructions to prepare Internal Solution II used for simultaneous patch clamp electrophysiology experiments to probe circuit connectivity.

Proper citation: Allen Institute for Brain Science 2020. Electrode Internal Solution with Biocytin II. protocols.io https://dx.doi.org/10.17504/protocols.io.bctsiwne Copy   

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Authors: Caroline Green
Summary: Adipose tissue, especially white adipose tissue (WAT), has been shown to be associated with endocrine and organ inflammation in addition to energy storage. Extraction and analysis of proteins from adipose tissue is increasingly important for understanding many physiological/pathological conditions. However, due to the high fat and low protein content in white adipose tissue (WAT) and brown adipose tissue (BAT), it is technically challenging.It is well known that water-oil emulsions in biological samples are the most difficult to separate, and the pore-sized centrifugal column with unique surface properties and the optimized surfactant-free buffer system can quickly and efficiently emulsify water oil from adipose tissue homogenate. The extraction buffer is lower than the oil freezing point in the adipose tissue, and the adipose tissue homogenate can quickly separate the aqueous phase and the oil phase through the centrifugal column, and the total protein in the tissue is not lost. Protein yields can reach 2-3 mg/ml, which is much higher than other methods.Collected by Creative BioMart

Proper citation: Caroline Green 2019. Total protein extraction from adipose tissue and cells. protocols.io https://dx.doi.org/10.17504/protocols.io.wurfev6 Copy   

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Authors:
Group: UCSC BME 22L

Proper citation: 2020. Protcol 1: PCR Dry Lab. protocols.io https:// Copy   

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Authors: Andrija S. Grustam, Nasuh Buyukkaramikli, Ron Koymans, Hubertus J.M. Vrijhoef, Johan L. Severens
Summary: Figure 1 - Single loop Monte Carlo scheme for computing overall individual Expected Value of Perfect Information (EVPI). andFigure 2 - Sampling algorithm for the calculation of Expected Value of Partially Perfect Information (EVPPI) with double loop.

Proper citation: Andrija S. Grustam, Nasuh Buyukkaramikli, Ron Koymans, Hubertus J.M. Vrijhoef, Johan L. Severens 2019. Value of Information in Telehealth for Chronic Heart Failure. protocols.io https://dx.doi.org/10.17504/protocols.io.xqdfms6 Copy   

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Authors: Allen Institute for Brain Science
Group: BICCN, Allen Institute for Brain Science
Summary: Quant-iT™ PicoGreen® dsDNA Assay is used for detection and quantitation of double stranded DNA products. Note: Research reported in this publication was supported by the National Institute Of Mental Health of the National Institutes of Health under Award Number U19MH114830. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Proper citation: Allen Institute for Brain Science 2020. 384 Well PicoGreen. protocols.io https://dx.doi.org/10.17504/protocols.io.bezejf3e Copy   

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Authors: Stemcell Technologies
Group: STEMCELL Technologies
Summary: Preparing a single cell suspension from a frozen starting sample is critical for optimizing cell isolations by avoiding additional cell loss and enabling the maximum labeling of target cells. Performing cell separation on clumpy cell samples can result in lower recovery and may interfere with proper labeling of the target cells. Samples may sometimes appear "clumpy" when they have been exposed to repeated freeze/thaw cycles or enzymatic tissue dissociation. These cell clumps occur because environmental stresses can accelerate the rate of cell death within the sample, resulting in the release of "sticky" DNA molecules from the dying cells that can clump neighboring cells together. Adding the endonuclease deoxyribonuclease I (DNase I) into your sample can minimize the presence of free-floating DNA fragments and cell clumps. This protocol describes a method to harvest cells and prepare a clump-free single cell suspension from a frozen cell sample prior to performing cell separation.

Proper citation: Stemcell Technologies 2019. How to Prepare a Single Cell Suspension from a Frozen Cell Sample. protocols.io https://dx.doi.org/10.17504/protocols.io.4umgwu6 Copy   

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Authors: Matt Sullivan Lab
Group: VERVE Net, Sullivan Lab
Summary: Must be made fresh before experiment because of the Sucrose. For 20 mL solutions.

Proper citation: Matt Sullivan Lab 2017. Lysis Buffer (20 mL). protocols.io https://dx.doi.org/10.17504/protocols.io.c4gytv Copy   

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