Pdf Slop99-Agar-Preparation-Without

Total Page：16

File Type：pdf, Size：1020Kb

Soy Fresh, Soy Clean Standard Laboratory Operating Procedure #99 Agar Preparation without an Autoclave Laboratory: Biotechnology Location: Ag Biotech Academy SOP prepared by: Rachel Sanders Last Revision: 05/12/16 General: This procedure is for the aseptic preparation of agar plates. The powdered agar medium bottle will have directions for preparing 1 L of agar. The fask that you are using needs to be at least twice the size of the amount of agar that you are preparing. The protocol listed below is for the preparation of 500 ml of agar in a 1000 ml Erlenmeyer fask. Click here for additional sterilization methods. Flinn Scientifc has a video with another method at their website. Log onto finnsci.com. click on Teacher Resources across the top. Click on Teacher Resource Videos at the bottom of the page. Click on Minute Videos. Click on Biology Videos. Select “Sterilizing without an Autoclave.”‎ Safety: PPE—eye protection, hot gloves, aprons MATERIALS: powdered agar medium Petri dishes distilled water stir rod cotton plug hot plate 1 L Erlenmeyer fask autoclave graduated cylinder balance hot gloves PROCEDURE: 1. The powdered agar medium bottle will have directions for preparing 1 L of agar. Divide the number of grams of powdered agar needed by 2 for the preparation of 500 ml. Measure out the powdered agar medium using a balance. 2. Measure 500 ml of distilled water into a graduated cylinder. 3. Pour the distilled water into the 1 L fask. 4. Pour 11.5g of the agar into the fask. 5. Swirl the fask to mix and add a magnetic stir bar. 6. Cover the fask with foil and heat with repeated stirring on the hot plate on high until the liquid is just boiling. Agar will boil over quickly. 7. Simmer the liquid for 30 minutes on low heat. 8. Allow the agar medium to cool to about 50o C before pouring into agar plates. 9. Pour the agar into the bottom of the Petri dish until 1/3 to 1/2 full. 10. Place the lid on the petri dish. 11. Agar plates sterilized in this manner are good for one to two days. 12. Option: if only need to make small batch of 4 plates, use 125ml of distilled water in step 2 and pour into a 250ml fask. Then for step 4, weigh out 5.75g of agar and add to fask. .

Copy Link

Recommended publications

Microbiology Laboratory Exercises Third Edition 2020
MICROBIOLOGY Laboratory Exercises Third Edition Keddis & Rauschenbach 2020 Photo Credits (in order of contribution): Diane Davis, Ines Rauschenbach & Ramaydalis Keddis Acknowledgements: Many thanks to those in the Department of Biochemistry and Microbiology, Rutgers University, who have through the years inspired our enthusiasm for the science and teaching of microbiology, with special thanks to Diane Davis, Douglas Eveleigh and Max Häggblom. Safety: The experiments included in this manual have been deemed safe by the authors when all necessary safety precautions are met. The authors recommend maintaining biosafety level 2 in the laboratory setting and using risk level 1 organisms for all exercises. License: This work is licensed under a Creative Commons Attribution- NonCommercial-NoDerivatives 4.0 International License Microbiology Laboratory Exercises Third Edition 2020 Ramaydalis Keddis, Ph.D. Ines Rauschenbach, Ph.D. Department of Biochemistry and Microbiology Rutgers, The State University of New Jersey CONTENTS PAGE Introduction Schedule ii Best Laboratory Practices Iii Working in a Microbiology Laboratory iv Exercises Preparation of a Culture Medium 1 Culturing and Handling Microorganisms 3 Isolation of a Pure Culture 5 Counting Bacterial Populations 8 Controlling Microorganisms 10 Disinfectants 10 Antimicrobial Agents: Susceptibility Testing 12 Hand Washing 14 The Lethal Effects of Ultraviolet Light 15 Selection of Fungi from Air 17 Microscopy 21 Morphology and Staining of Bacteria 26 Microbial Metabolism 30 Enzyme Assay 32 Metabolic
[Show full text]
Page 1 of 27 the Diagram Shows Two Thermometers. the Bulb of Each
Q1. The diagram shows two thermometers. The bulb of each thermometer is covered with a piece of wet cotton wool. One of the thermometers is placed in the draught from a fan. The graph shows how the temperature of each thermometer changes with time. (a) Which of the graph lines, A or B, shows the temperature of the thermometer placed in the draught? Write the correct answer in the box. Page 1 of 27 Explain, in terms of evaporation, the reason for your answer. ___________________________________________________________________ ___________________________________________________________________ ___________________________________________________________________ ___________________________________________________________________ ___________________________________________________________________ ___________________________________________________________________ (3) (b) A wet towel spread out and hung outside on a day without wind dries faster than an identical wet towel left rolled up in a plastic bag. Explain why. ___________________________________________________________________ ___________________________________________________________________ ___________________________________________________________________ ___________________________________________________________________ (2) (Total 5 marks) Q2. The picture shows a person taking a hot shower. (a) When a person uses the shower the mirror gets misty. Why? ___________________________________________________________________ ___________________________________________________________________
[Show full text]
Autoclave Quick Guide
Office of Biological Safety Autoclave Operation Quick Reference Guide Training: ALL users MUST undergo documented training for operation of the autoclave. Record each use of the autoclave in a log: Date, User ID, cycle type and nature of material in load Personal Protective Equipment (PPE) needed: ¾ Wear eye protection, lab coat, gloves along with heat resistant gloves. Rubberized apron, sleeve guards and face shield are recommended when autoclave is hot or splash risk is present. Hints and Precautions ¾ Become familiar with the manufacturer’s operations manual of your autoclave model(s). ¾ Plastics used for autoclaving MUST be labeled as autoclavable – otherwise the plastic will melt. ¾ Waste bags to be autoclaved must be loosely packed and not more than 2/3 filled. Steam must be able to penetrate to all contents of the bag. ¾ Sharps or pointed hard objects should not be placed directly into an autoclave bag; a thicker or rigid container must be used (such as a sharps container). ¾ Avoid overfilling an autoclave with loads or allowing a load to contact the chamber walls. ¾ Transferring waste contents from an overfilled bag to another bag should be avoided! This practice can lead to injury and/or exposure to contaminants. ¾ Do not leave an autoclave operating unattended for long periods of time; operation should be monitored periodically during a cycle in case of failure. ¾ Never autoclave solvents, combustible, volatile, flammable, radioactive or corrosive materials (e.g. ethanol, methanol, acids, bases, phenol) ¾ Remove extraneous items and combustible materials from around the autoclave exterior. Loading ¾ Follow manufacturer’s loading instructions for your autoclave model ¾ Transport loads on a cart and in secondary containers to reduce spills ¾ Clean item/container sterilization Loosen caps or lids to avoid dangerous pressure build-up during cycle Place containers in a tray and load the tray into the autoclave – this is easier to load and unload and will catch spills.
[Show full text]
Pouring Plates from Prepared Bottled Media
Pouring Plates from Prepared Bottled Media Primary Hazard Warning Never purchase living specimens without having a disposition strategy in place. When pouring bottles, agar is HOT! Burning can occur. Always handle hot agar bottles with heat-protective gloves. For added protection wear latex or nitrile gloves when working with bacteria, and always wash hands before and after with hot water and soap. Availability Agar is available for purchase year round. Information • Storage: Bottled agar can be stored at room temperature for about six months unless otherwise specified. Never put agar in the freezer. It will cause the agar to breakdown and become unusable. To prevent contamination keep all bottles and Petri dishes sealed until ready to use. • Pouring Plates • Materials Needed: • Draft-free enclosure or Laminar flow hood • 70% isopropyl alcohol • Petri dishes • Microwave or hot water bath or autoclave 1. Melt the agar using one of the following methods: a) Autoclave: Loosen the cap on the agar bottle and autoclave the bottle at 15 psi for five minutes. While wearing heat-protective gloves, carefully remove the hot bottle and let it cool to between 75–55°C before pouring. This takes approximately 15 minutes. b)Water Bath: Loosen the cap on the agar bottle and place it into a water bath. Water temperature should remain at around 100°C. Leave it in the water bath until the agar is completely melted. While wearing heat- protective gloves, carefully remove the hot bottle and let it cool to between 75–55°C before pouring. c) Microwave: Loosen the cap on the agar bottle before microwaving.
[Show full text]
Biohazardous Waste Handling for Eastern Kentucky University May 2018
Biohazardous Waste Handling For Eastern Kentucky University May 2018 Approved by The University Laboratory Safety Biohazard Subcommittee 1 Table of Contents Page Introduction 3-4 Definitions 5-6 Responsibilities and roles 7-9 Main body of document 10-17 Appendix 1 18-22 Autoclave procedures in SB 18-20 Autoclave procedures in Disney 218 21-22 Appendix 2 copy of IACUC form H: Use of hazardous agents 23 2 3 Introduction This document has been prepared to provide guidance to Eastern Kentucky University employees and students in the use and disposal of biohazardous materials in compliance with regulatory requirements. For this document “biohazardous waste” is defined as any discarded material which might include infectious laboratory materials or agents regulated by federal, state, and local authorities. At a minimum, the following categories should be considered as biohazardous waste materials: 1. Cultures and stocks of infectious biological agents, including laboratory waste, discarded live or attenuated viruses or related agents, culture dishes and other laboratory supplies used in the production or use of these agents, and any other related devices. 2. Human blood is a biohazardous waste. Human blood should treated in accordance with the “Eastern Kentucky University Bloodborne Pathogens Exposure Control Plan”. This plan can be found at the Risk Management Insurance/ Environmental Health and Safety website https://ehsrmi.eku.edu/occupational-safety. 3. Sharps: These are defined as needles, syringes, scalpels, etc., as well as any object sharp enough to puncture the skin (i.e. microscope slides, cover slips) that is used in the laboratories that could possibly come in contact with material that may be considered biohazardous waste.
[Show full text]
Microlab® STAR™
Microlab ® STAR™ Microlab ® STAR ™ AUTOMATED WORKFLOW SOLUTIONS CENTERED AROUND YOUR ASSAY The STAR combines Hamilton's patented pipetting technology including precise lock-and-key tip attachment, unrivaled liquid level detection, and comprehensive volume ranges to create flexible liquid handling workstations. Available in three base platform sizes, the STAR portfolio incorporates countless options to automate your workflows. Hamilton Robotics has also partnered with top leaders in the biotechnology industry to provide Standard Solutions based on commonly automated applications. Offering ready-to-start protocols for a variety of applications such as NGS, ELISA, and forensic assays, our Standard Solutions provide a faster way to automate your processes. 2 1 PATENTED TECHNOLOGY The STAR utilizes Hamilton’s proprietary Compressed O-Ring Expansion (CO-RE®) technology. CO-RE minimizes the production of aerosols and allows disposable tips or washable, steel needles to be used on channels in the same run. 2 MULTI-FUNCTIONAL ARM Our technology offers high pipetting accuracy and precision, from sub-microliter to large volumes, using Independent Channels and/or the Multi-Probe Head (MPH). Labware transportation is possible with the iSWAP® or CO-RE Grippers. The STAR can incorporate a camera, tube transportation, and other channel tools on a single arm. Comprehensive pipetting range: ■■■0.5 μL to 1 mL using the 1 mL Independent Channel ■■■50 μL to 5 mL using the 5 mL Independent Channel ■■■1 μL to 1 mL using the CO-RE 96 MPH ■■■0.1 μL to 50 μL using the CO-RE 384 MPH 3 FLEXIBLE SETUP The high-capacity deck is customized specific to your workflow, accommodating a wide range of labware and automated devices that can easily be exchanged to support multiple assays on one platform.
[Show full text]
The Eppendorf Mechanical Pipettes Research® Plus and Reference 2 – Fully Autoclavable, Easy Adjustment, Quick and Simple Maintenance
APPLICATION NOTE No. 198 The Eppendorf Mechanical Pipettes Research® plus and Reference 2 – Fully Autoclavable, Easy Adjustment, Quick and Simple Maintenance Kornelia Ewald, Ulrike Gast, Eppendorf AG, Hamburg, Germany Abstract Research plus and Reference 2 pipettes are easy to clean safe to perform. This Application Note demonstrates the and maintain. Hence, routine maintenance can be easily special features of maintaining and cleaning Research performed by the user, saving valuable time. Readjust- plus pipettes and provides detailed information on ment of the pipette for liquids whose physical properties changing the pipette‘s adjustment. diff er signifi cantly from those of water is also easy and Introduction Research plus and Reference 2 are manual pipettes with Adjustment for speciﬁ c liquids or altitude diff erent features and functional principles (e.g. one-button / During production, piston stroke pipettes are adjusted two-button operation). Fulfi lling the highest requirements, to distilled water under certifi ed measuring conditions. both pipettes have equal functionalities in terms of decon- To indicate the adjustment, all Research plus pipettes carry tamination, maintenance and adjustment. In order to meet an adjustment seal (Fig. 1). If necessary, adjustment for the high requirements of a modern laboratory, high quality specifi c liquids or for altitude can be carried out easily by piston stroke pipettes should be partly or fully autoclavable the user. The red adjustment seal, which is applied to the as well as UV resistant. adjustment opening following an adjustment, serves to The Eppendorf Research plus and Reference 2 pipettes can visualize a change of adjustment settings (Fig. 1). be decontaminated either by UV light or by autoclaving the entire instrument.
[Show full text]
Elara11 Autoclave Quick Installation Guide
OVER YEARS S ince 1925 Elara11 Autoclave Quick Installation Guide 1. Examine the outer carton and autoclave for any signs of damage. Immediately notify your dealer or Tuttnauer USA of any signs of damage. 2. To avoid injuries, lifting and carrying should be done by two people. • Lifting straps have been installed for your convenience. Lifting straps are for one time use only and should be removed and discarded after initial set up. 3. Place the sterilizer on a rigid level surface. The counter top or stand must be able to support 275lbs lbs Minimum 24”depth 4. The minimum depth of the counter top needs to be 24 inches. counter top required 5. It is mandatory to leave a minimum of 2” clearance between the back of the Elara11 and the wall. If located in a cabinet, the rear panel of the cabinet must be removed for proper air circulation. Failure to provide the needed clearance will result in failed cycles. 6. Side clearances should be a minimum of 2”. 7. Make sure all the feet are on the autoclave and none of them have been lost. 8. Connect the power cord to the socket on the rear of the autoclave; then plug it into the supply outlet. Power switch a. This unit requires a 230 volt 1 or 2 phase 15A supply. b. The acceptable operating voltage range is 220 to 235 volts. c. The installation of a Buck/Boost transformer (0.5KVA) may be required to meet the acceptable operating voltage. d. The supply outlet must be a properly grounded outlet.
[Show full text]
Cleaning and Decontamination Guide for Sartorius Pipettes
Operating Manual Cleaning and Decontamination Guide for Sartorius Pipettes Version 1 / 2018 / 11 Contents Contents 1 Cleaning and Decontamination in Pipette Maintenance . 4 2 Cleaning Guide . 5 2 .1 Cleaning the outer surface of the pipette (daily) . 5 2 .2 Cleaning the lower part of the pipette (every three months) . 5 3 Application-Specific Cleaning and Decontamination of Pipettes . 10 3 .1 When cleaning Sartorius pipettes for specific applications . 10 3 .2 Cleaning and decontamination reagent quick-guide . 11 3 .3 Cleaning agents and their activities . 11 3 .4 Cleaning procedures . 12 3 .5 Decontamination agents and their activities . 12 3 .6 Decontamination procedures . 13 4 Autoclaving Guide . 14 4 .1 Mechanical pipettes . 14 4 .2 Electronic pipettes . 14 4 .3 Autoclaving quick-guide . 15 4 .4 Drying of pipettes . 15 5 Summary of Pipette Parts . 16 5 .1 Mechanical pipettes parts and materials . 16 5 .1 .1 Tacta® mechanical pipette . 16 5 .1 .2 mLine® mechanical pipette . 17 5 .1 .3 Proline® Plus mechanical pipette . 17 5 .1 .4 Proline® mechanical pipette . 18 5 .2 Electronic pipette parts and materials: Picus and Picus® NxT . 18 6 Contact . 19 6 .1 Further information about pipette cleaning, decontamination and maintenance . 19 6 .2 Pipette maintenance, servicing or calibration . 19 Cleaning and Decontamination Guide for Sartorius Pipettes 3 Cleaning and Decontamination in Pipette Maintenance 1 Cleaning and Decontamination in Pipette Maintenance Pipettes are precision instruments whose performance can be significantly impacted if not adequately maintained . To maintain the level of purity that is critical in many laboratory applications, adequate cleaning and decontamination of pipettes is necessary .
[Show full text]
Evobot: an Open-Source, Modular, Liquid Handling Robot for Scientific Experiments
applied sciences Article EvoBot: An Open-Source, Modular, Liquid Handling Robot for Scientific Experiments Andres Faiña 1,* , Brian Nejati 2 and Kasper Stoy 1 1 Robots, Evolution and Art Lab (REAL), Department of Computer Science, IT University of Copenhagen, 2300 Copenhagen, Denmark; [email protected] 2 Department of Mechanical and Industrial Engineering, University of Toronto, Toronto, ON M5S 1A1, Canada; His work was carried out while he was employed at ITU; [email protected] * Correspondence: [email protected]; Tel.: +45-7218-5258 Received: 13 October 2019; Accepted: 17 January 2020; Published: 23 January 2020 Featured Application: Our modular liquid handling robot can be applied to automate scientific experiments in research labs, where tasks change often, and to carry out new kinds of experiments that cannot be done manually. Abstract: Commercial liquid handling robots are rarely appropriate when tasks change often, which is the case in the early stages of biochemical research. In order to address it, we have developed EvoBot, a liquid handling robot, which is open-source and employs a modular design. The combination of an open-source and a modular design is particularly powerful because functionality is divided into modules with simple, well-defined interfaces, hence customisation of modules is possible without detailed knowledge of the entire system. Furthermore, the modular design allows end-users to only produce and assemble the modules that are relevant for their specific application. Hence, time and money are not wasted on functionality that is not needed. Finally, modules can easily be reused. In this paper, we describe the EvoBot modular design and through scientific experiments such as basic liquid handling, nurturing of microbial fuel cells, and droplet chemotaxis experiments document how functionality is increased one module at a time with a significant amount of reuse.
[Show full text]
The Effect of Autoclave Resterilisation on Polyester Vascular Grafts
CORE Metadata, citation and similar papers at core.ac.uk Provided by Elsevier - Publisher Connector Eur J Vasc Endovasc Surg 18, 386±390 (1999) Article No. ejvs.1999.0891 The Effect of Autoclave Resterilisation on Polyester Vascular Grafts G. Riepe∗1, M. S. Whiteley2, A. Wente1, A. Rogge1, A. SchroÈ der1, R. B. Galland2 and H. Imig1 1Department of General, Vascular and Thoracic Surgery, General Hospital of Hamburg±Harburg, Germany; 2Department of Surgery, Royal Berkshire Hospital, Reading, U.K. Objectives: polyester grafts are expensive, single-use items. Some manufacturers of uncoated, woven grafts include instructions for autoclave resterilisation to be performed at the surgeon's own request. Others warn against such manipulation. Theoretically, the glass transition point of polyester at 70±80°C and the possible acceleration of hydrolysis suggest that autoclave resterilisation at 135°C might be a problem. Materials and methods: a DeBakey Soft Woven Dacron Vascular Prosthesis (Bard) and a Woven Double Velour Dacron Graft (Meadox) were autoclave-resterilised 0 to 20 times, having been weighed before and after sterilisation. Tactile testing was performed. Mechanical properties were examined by probe puncture and single-®lament testing, the surface was examined by scanning electron microscopy and the degree of hydrolysis by infra-red spectroscopy. Results: tactile testing revealed a change of feeling with increasing cycles of resterilisation. Investigation of weight, textile strength, single-®lament strength, electron microscopy of the surface and infra-red spectroscopy showed no change of the material. Conclusions: changes felt are presumably a surface phenomenon, not measurably affecting strength or chemistry of material after autoclave resterilisation.
[Show full text]
Transforming Plasmid DNA Into Electrocompetent Cells
Transforming plasmid DNA into electrocompetent cells 1. Clean and dry electroporation cuvettes throroughly on the cuvette washer. Chill on ice and allow to air dry. Use one cuvette for each DNA sample you are transforming. 2. Dialyze your DNA sample(s) using a nitrocellulose filter and DI water. • Fill a Petri dish with DI water. • Place a single nitrocellulose filter paper on the surface of the water – shiny side up. If you are dialyzing more than one sample, it helps to cut a small notch in the paper to serve as a “key”. This helps to identify/keep track of your samples. • Spot DNA samples onto the surface of the paper. Space out the samples a little bit because they will expand in volume as they dialyze. • Dialyze your sample for 10-15 minutes. *This step is particularly important if the DNA has undergone any previous manipulations that have introduced salts (adding buffers for ligation, restriction enzyme digestions, etc.). Too much salt in the DNA will cause your sample to arc when electroporating. This will kill the cells and significantly reduce the efficiency of transformation. If your sample arcs, it may still be worth while to plate your cells. However, it is likely that you will get very few (if any) colonies. You should repeat this sample. Main factors that can cause arcing - too much salt in your DNA, water on the outside of the cuvette, oil on the outside of the cuvette from handling it too much without gloves, too much salt in the cells. 3. Thaw electrocompetent cells on ice.
[Show full text]