Q&A Aug 11, 2026

FITC labeling of S.pneumoniae Protocol        Aim: Labeling of bacteria with FITC Requirements Fluorescein isothiocyanate (FITC, isomer I; Sigma Chemicals Co)DMSO PBS1x or Bicarbonate buffer (0.1M NaHCO3) pH ~8.5;  84mg in 10ml MQ Assay                                    Prepare 10 mg/ml FITC solution in DMSO (10mg in 1ml DMSO)Grow bacteria in 10 ml THY (~0.6 OD550) Transfer to 15 ml falcon tube and harvest bacteria by centrifugation at 3500 rpm, 10 min.Wash bacteria once with 5 ml bicarbonate b...

Q&A Aug 10, 2026

 Materials1) TCEP HCl, Cat#75259, Sigma-Aldrich2) P6 spin-column (Cat# 7326227, BioRad)3) Phosphate Buffered Saline without calcium chloride and magnesium chloride, Cat#D8537, Sigma-Aldrich)4) Alexa 594 maleimide (Cat#A10256, Invitrogen)5) Superdex 200 ProceedureStart with an eppendorf tube containing a solution of nanobody (100 micrograms/0.1 mL), add 1.5 microL of tris-(2-carboxyethyl)phosphine (1 M TCEP HCl, Cat#75259, Sigma-Aldrich). The final concentration of TCEP is 15 mM.Shake the mixture...

Q&A Aug 9, 2026

Title: UVB‑induced senescence of HaCaT cellsAuthors:Jiayu Liu (Clarity0103@163.com)Xiaobo Wang(wxb4320062@163.com)School of Basic Medicine, Dali University, Dali, 671000, Yunnan, China1. AbstractThis protocol describes an in vitro model of cellular senescence induced by ultraviolet B (UVB) irradiation in human immortalized keratinocytes (HaCaT cells). By applying two consecutive daily exposures of different UVB doses, the method mimics  photo‑damage and is suitable for studying skin photoaging a...

Q&A Aug 7, 2026

I. Protein Expression The protein expression protocol described below can be used for all DDX-23, MAB-10, and fluorescent proteins used in the study. For DDX-23 and MAB-10 it is best to produce 2 liters of main culture to obtain a useful amount of protein. For the fluorescent proteins alone, half a liter of main culture is sufficient. Materials and ReagentsShuffle T7 competent E. coli cells (New England Biolabs)LB BrothAmpicillinFlasks (preferably baffled)IPTGCulture tubes15 mL tubesCentrifuge b...

Q&A Aug 7, 2026

One-Step Phage Growth CurveA. Preparation of the bacterial hostGrow the bacterial host culture at 37 °C with shaking until the exponential growth phase is reached (OD600=0.5–0.6).Adjust or estimate the bacterial concentration to approximately 108 CFU/mL.B. Phage infectionMix 0.9 mL of the bacterial suspension (approximately 108 CFU/mL) with 0.1 mL of phage suspension (approximately 107 PFU/mL) to obtain an MOI of 0.01.Incubate the phage–host mixture for 10 min at 37 °C to allow phage adsorption....

Q&A Aug 4, 2026

GCase Activity Assay Using PFB-FDGlu in SH-SY5Y Cells (96-Well Plate Format)Priya Thakur1, Han Seok Ko11Neuroregeneration and Stem Cell Programs, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA; Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. PrinciplePFB-FDGlu is a cell-permeable fluorogenic substrate that is hydrolyzed by lysosomal GCase in a live cell to release a fluorescent product. Fluores...

Q&A Jul 23, 2026

An Efficient Parallel Single-Primer PCR Method for Site-Directed Mutagenesis Abstract Regulation of gene expression by histone modifications is a fundamental process in cells. Dysregulation of epigenetic regulators, either by altered expression or activity, is a hallmark of many cancers and contributes to the initiation and progression of tumors. Notably, cellular metabolism is closely intertwined with epigenetic regulation, as several metabolites directly influence the epigenetic landscape. Her...

Q&A Jul 22, 2026

SCENITH PROTOCOLStandard Operating Procedure for Single-Cell Energetic Metabolism Profiling by Flow CytometryProtocol Name:SCENITH Protocol (Single-Cell Energetic Metabolism Profiling by Translation Inhibition)Target Application:Intracellular Flow Cytometry / Metabolic ProfilingIncubation Conditions:37°C, 5% CO2 / 4°C / Room Temperature (RT) 1. Step-by-Step Execution ProcedureStep #Procedure / ActionStep 1Pre-heat the three metabolic drugs (C, DG, O) and puromycin at 37°CStep 2Label Eppendorf tu...

Q&A Jul 17, 2026

OverviewThis assay quantifies phagocytic activity of Drosophila larval hemocytes using fluorescent bacterial bioparticles and flow cytometry. Hemocytes are identified/gated using Hml-Gal4, UAS-GFP, and phagocytosis is assessed by uptake of Alexa Fluor 488–labeled bioparticles.Step 1: Definition of Hemocyte Gating Using Hml-Gal4, UAS-GFPThis step is performed periodically to establish robust hemocyte gates and instrument settings on the CytoFLEX using a Hml-Gal4, UAS-GFP line, enabling subsequent...

Q&A Jul 15, 2026

A SIMPLE AND REPRODUCIBLE METHOD FOR EXTRACTION OF VIABLE SEEDS FROM RIPE TOMATO FRUITSAnjan Barman*Department of BiotechnologyPandu College, Maligaon, Guwahati-12, Assam, India*Address for Correspondence: E-mail: anjbth@gmail.comABSTRACT:Tomato seeds are harvested from mature and ripe fruits that involve extraction and separation of seeds from plant materials. Mechanical and chemical methods for separation of tomato seeds from plant materials are in regular practice. Here, a very simple approac...