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GEN Protocols

Translational Medicine

Translational Medicine

Protocol

Short Communication: Update in Natural Antiretroviral Resistance-Associated Mutations Among HIV Type 2 Variants and Discrepancies Across HIV Type 2 Resistance Interpretation Tools

HIV variants carry natural polymorphisms related to drug resistance (R-markers) fixed during viral evolution in the absence of antiretroviral therapy (ART) that may impact on drug susceptibility and resistance pathways. We aimed to identify the HIV type 2 (HIV-2) variant-specific R-markers at Pol in... More


Protocol

Short Communication: Coronavirus Disease 19 Among People Living with HIV in Western India: An Observational Cohort Study

A retrospective cohort study was conducted to assess clinical characteristics and outcomes of coronavirus disease-19 (COVID-19) among people living with HIV (PLHIV) in western India. Out of 86 PLHIV with COVID-19 illness, 19.7% had severe/critical illness and 6 (6.9%) individuals died. Median (inter... More


Protocol

Short Communication: Higher Tenofovir Concentrations in Hair Are Associated with Decreases in Viral Load and Not Self-Reported Adherence in HIV-Infected Adolescents with Second-Line Virological Treatment Failure

Objective methods of measuring antiretroviral adherence are limited. We assessed the relationship between tenofovir disoproxil fumarate (TDF) hair concentrations, self-reported adherence, and virological outcomes in HIV-infected adolescents in Harare, Zimbabwe. HIV-infected adolescents on atazanavir... More


Protocol

Continuous Glucose Monitoring and Glycemic Control in Young Adults with Type 1 Diabetes: Benefit for Even the Simplest Insulin Administration Methods

The impact of continuous glucose monitoring (CGM) on glycemic control in young adults with type 1 diabetes (T1D) is controversial. Data from 888 young adults with T1D were reviewed (ages 18–30 years, 52% female, glycated hemoglobin [HbA1c] 8.1% ± 1.5%). Prescription of CGM was recorded for 54% of yo... More


Protocol

School Nurse Perspectives on Do-It-Yourself Automated Pancreas Systems in the School Setting

Do-it-yourself (DIY) artificial pancreas systems (APSs) are gaining popularity among children with type 1 diabetes. Little is known about how school systems provide care for children who use DIY APSs, and available guidance for schools is limited. This study explored school staff perspectives on DIY... More


Protocol

Correlates of Continuous Glucose Monitoring Use Trajectories in Children and Adolescents with Type 1 Diabetes

The goal of this study was to characterize trajectories of continuous glucose monitoring (CGM) use in youth 5–12 weeks after starting CGM and examine what factors differentiate between the trajectory groups. Parent–youth dyads completed assessments before starting CGM. Days of CGM use between weeks ... More


Protocol

Continuous Glucose Monitoring Time-in-Range and HbA1c Targets in Pregnant Women with Type 1 Diabetes

The CONCEPTT trial compared real-time Continuous Glucose Monitoring (RT-CGM) to capillary glucose monitoring in pregnant women with type 1 diabetes. We analyzed CGM and glycated hemoglobin (HbA1c) measures in first (n = 221), second (n = 197), and third (n = 172) trimesters, aiming to examine target... More


Protocol

Insulin Metrics: Need for Development of Consensus Standards for Reporting of Insulin Dosing Data

The advent of connected insulin pens will generate an avalanche of digital insulin data, especially in the context of prandial- and multiple daily injection-insulin regimens. There is a need for the diabetes community to develop standards for such data, analogous to what has been achieved using the ... More


Protocol

Predictive Low-Glucose Suspend Necessitates Less Carbohydrate Supplementation to Rescue Hypoglycemia: Need to Revisit Current Hypoglycemia Treatment Guidelines

Current guidelines recommend 15–20 g of carbohydrate (CHO) for treatment of mild to moderate hypoglycemia. However, these guidelines do not account for reduced insulin during suspensions with predictive low-glucose suspend (PLGS). We assessed insulin suspensions, hypoglycemic events, and CHO treatme... More


Protocol

Zebrafish Tumor Graft Transplantation to Grow Tumors In Vivo That Engraft Poorly as Single Cell Suspensions

Angiosarcoma is a clinically aggressive tumor with a high rate of mortality. It can arise in vascular or lymphatic tissues, involve any part of the body, and aggressively spread locally or metastasize. Angiosarcomas spontaneously develop in the tp53 deleted (tp53del/del) zebrafish mutant. However, e... More


Protocol

A Protocol for Qualifying DNA Variants Associated with Complex Diseases like Ehlers-Danlos syndrome

A protocol for DNA variant qualification adds medical perspective to consensus grading based on impact of the DNA change on its encoded RNA/protein and its prevalence in normal people versus those with the implicated disease. This clinically familiar 1-4+ qualification can encourage physician accept... More


Protocol

Investigating the increased lifespan in C. elegans daf-2 mutants by 4D-Lipidomics™

The small nematode Caenorhabditis elegans is one of the premier biomedical model organisms and employed in many aspects of basic and applied science.... More


Protocol

4D-Lipidomics™ workflow for increased throughput

Lipid profiling from complex lipid extracts can be a challenging and time consuming task. The high complexity of samples and co-elution of isobaric or isomeric compounds complicate the confident annotation of lipids. The presented 4D-Lipidomics workflow simplifies and streamlines the annotation and ... More


Protocol

Rapid metabolic phenotyping in SARS-CoV-2 patients using the impact II QTOF MS system

The impact II QTOF system meets the critical elements of sensitivity, selectivity, accuracy, and precision for targeted metabolite analysis. The continuous collection of both MS and MS/MS data provides an exponentially deeper and broader pool of data valuable for subsequent retrospective discovery o... More


Protocol

Removal of rare undifferentiated induced pluripotent stem cells (iPSCs) by multi-step negative sorting

In the field of regenerative medicine involving ES or iPS cells, differentiated cells are often cultured to tissues and implanted to patients. However, there is an unneglectable problem that some remaining undifferentiated cells in the implanted tissue may cause tumors. Therefore, a complete removal... More


Protocol

Sorting of water-in-oil emulsions with bacteria using a fluorescent nucleic acid probe

To isolate water-in-oil emulsion droplets containing bacteria from those that do not contain bacteriawe have developed a simple and reliable selective sorting method that allows bacteria-containing droplets to be isolated on our microfluidic chip cell sorter, On-chip Sort. Bacteria-containing drople... More


Protocol

Cell culture CO2 incubator decontamination process using hydrogen peroxide vapor for highly regulated and general cell culture protocols

The value of the laboratory cell culture incubator used in highly regulated research and clinical protocols is directly related to the proportion of incubator uptime vs. downtime in applications where frequent interior chamber decontamination is required or desired. The need for interior decontamina... More


Protocol

Isolation of Mononuclear Cell Fractions Using Your Sepax™ System

The purpose of this protocol is to provide instructions on how to isolate mononuclear cell fractions of apheresis product, bone marrow, or umbilical cord blood using the Sepax™ system with initial volumes of 30 to 120 mL and selectable final volumes of 8-20 mL or 45 mL. This protocol applies to anyo... More


Protocol

How to run cryopreservation protocols using a VIA Freeze™ system

The purpose of this protocol is to give the reader a comprehensive understanding of how to set up and operate the Cytiva VIA Freeze™ Uno, VIA Freeze™ Duo, and VIA Freeze™ Quad units.This protocol applies to anyone performing a procedure with the VIA Freeze™ units. Once training on this document is c... More


Protocol

Streamlined High Throughput Drug Screening with PrimeSurface® White 384 well 3D plates

Using PrimeSurface® white plate, cancer spheroid cells can be formed and tested for drug screening using luminescence viability assay in the same well without the need for reagent transfer. This streamlined process shortens the experimental steps and time required in drug testing thus providing a ro... More


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Primer

Two Critical Steps to Success in Multicolor Flow Cytometry

Flow cytometry is a powerful tool to analyze and characterize cells based on their protein expression, most often those expressed on the cell surface. Multicolor flow cytometry uses multiple fluorochromes as primary or secondary antibody conjugates to label cells. The capacity for modern flow cytome... More


Primer

Expressing, Extracting and Purifying Membrane Proteins

Membrane proteins are key biological players involved in myriad essential cellular processes from molecular transport, cellular communication, and signal transduction to maintaining structures within cells and tissues. Despite their importance, accounting for 25% of all proteins in the human genome ... More


Primer

Unwanted Fungi: Detecting Microbial Cell Culture Contamination

Coming into the lab and spotting an obvious fungal growth, or the tell-tale signs of cloudy media, can be a stressful experience for researchers. When carrying out cell culture in the lab, contamination from microbial agents such as bacteria and fungus can occur and have disastrous effects on ongoin... More


Primer

Source and Elimination of Common Mass Spectrometry Contaminants

Sensitivity is a key attribute of mass spectrometers to allow quantification of minute amounts of analytes. Yet this sensitivity can be a conundrum. Myriad sources can contribute contaminants that can impact background and confound interpretation of results. An increased background signal can make i... More


Primer

Assessing qRT-PCR COVID-19 Testing Results: What Can Go Wrong?

COVID-19 is commonly diagnosed by quantitative fluorescence real-time reverse transcriptase-polymerase chain reaction (qRT-PCR) (1, 2). The test is the sequential action of two enzymes--an RNA-dependent DNA polymerase, also known as a reverse transcriptase, to copy the SARS-CoV-2 RNA into complement... More


Primer

Pathogen Detection Assays: Advantages, Disadvantages and Alternatives

Viral infections are difficult to diagnose based on the patient’s clinical presentation alone. A reliable and accurate diagnosis requires laboratory-based testing for the pathogen of interest. Such laboratory testing involves either the direct detection of viral antigens, or detection of virus-speci... More


Primer

Benefits of Using an Electronic Laboratory Notebook

Electronic laboratory notebooks (ELNs) were developed to keep track of experiments and data so that scientists could review their work in a single place and easily reproduce experiments. Many ELNs started out as electronic versions of paper lab notebooks. Now, the most advanced ELNs offer an extensi... More


Primer

Qualitative, Semi-quantitative, and Quantitative Immunodiagnostics for COVID-19

While qualitative reverse transcription polymerase chain reactions (RT-PCR) are used to diagnose COVID-19, serologic tests for antibodies specific to the SARS-CoV-2 virus are used to detect past infection. Most serologic assays are qualitative and semi-quantitative but quantitative serologic assays ... More


Primer

Saliva, Nasal Swabs, or Blood: Best Samples for Pathogen Detection

Most immunodiagnostic protocols and commercially available kits require serum, plasma, or blood samples. Collecting such samples requires the skills of a phlebotomist. For pathogen detection, particularly in largescale field studies, the collection of noninvasive samples such asnasal swabs, tears, o... More


Primer

Separating Blood into Cellular and Non-Cellular Components

A key diagnostic step in disease detection is analyzing blood components. Accurate separation of blood cells from plasma, which holds proteins and other constituents, is crucial in many assays.First introduced in the late 1980s, lateral flow devices, also referred to as lateral flow immunoassays, ar... More


Mixer

Organoids for Airway and Alveolar Disease Modeling

In this Mixer, we will discuss the development of respiratory airway and alveolar organoids derived from human iPSCs and how these are used in modeling respiratory diseases such as cystic fibrosis. Airway organoid models can be used for testing the therapeutic efficacy of candidate drugs for respira... More


Video

GEN Protocols Expert Exchanges: Tools and Techniques to Develop Therapies for Neurological Disorders

The human brain is an exquisitely complex organ, consisting of nearly one hundred billion neurons, an equivalent number of non-neuronal cells and trillions of synapses. Understanding the factors that play a role in the development of the structure and function of the brain is an essential step to de... More


Video

GEN Protocols Expert Exchanges: Patient-Derived Pancreatic Cancer Organoids Predict Response to Therapy

Pancreatic cancer accounts forthree percentof all cancers in the United States and is responsible for seven percent of all cancer deaths. Miniature organoids cloned from pancreatic tumors in patients are being grown in laboratories to conduct assays that can predict the effectiveness of chemotherapy... More


Video

GEN Protocols Expert Exchanges: Critical ALS Biomarkers of Neuroinflammation & Oxidative Stress

Biomarkers are essential tools for the development of therapeutics, clinical trial design and patient stratification. Biomarkers also help evaluate the efficacy of therapeutic intervention on disease progression or relapse. Yet, biomarker driven patient stratification, a hallmark in oncology trials,... More


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