MicroStation System, MicroLog Version 4.2 - DTU Systems Biology ...

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MicroPlates of aerobic bacteria can be read after 4-6 hours of incubation. If reactions are insufficient, read again at 16-24 hours. Preparing Samples 7. If the fluid level in the tips gets low, refill and continue dispensing until all wells are full. 8. Cover the MicroPlate with its lid. Incubating MicroPlates As soon as you dispense the suspension, incubate the MicroPlate using the appropriate temperature, atmosphere, and time conditions. Table 4-3 (page 4-15) and Appendix 2 will help you select the correct conditions. • For aerobes: Use a moist chamber during incubation to prevent evaporation. A plastic container with wet paper towels on the bottom should be sufficient. • For filamentous fungi: MicroPlates must be incubated in a closed container or in sealed plastic bags on trays. Do not add a source of moisture. Mycelial growth will overwhelm the MicroPlate and cause reaction crossover within the MicroPlate. Organism Type Temperature Atmosphere Incubation Time Gram Negative Non-Enteric 30° C Air 4-6 and 16-24 hours Gram Negative Enteric 35-37° C Air 4-6 and 16-24 hours Gram Negative Fastidious 35-37° C 6.5% CO2 4-6 and 16-24 hours Gram Positive Cocci Gram Positive Rods (nonspore forming) Gram Positive Rods (spore forming bacillus) 35-37° C or 30° C or 26° C (if required) 30° C or 55° C (if required) Anaerobes 35° C or 30° C or 26° C (if required) Air or 6.5% CO2 (if required) 4-6 and 16-24 hours Air 4-6 and 16-24 hours Anaerobic-H2 free 20-24 hours Yeasts 26° C Air 24, 48, or 72 hours Filamentous Fungi (and select yeast species) 26° C Air 24, 48, 72 or 96 hours TABLE 4-3: SELECTING CORRECT INCUBATING CONDITIONS MicroStation System/MicroLog User Guide Apr 07 Section 4 � Page 15
Preparing Samples Special Procedures for Incubating Anaerobic MicroPlates Caution! If you’re working with anaerobes: �Use anaerobically prepared media. �Inoculate MicroPlates in air. �After inoculation, wait 10 minutes before moving the MicroPlates into a hydrogenfree anaerobic jar. 1. In most cases, leave AN MicroPlates in their foil pouch until immediately before inoculation. The only exception is if you presume your organism is either Bacteroides or Prevotella. If your gramnegative rod is resistant to kanamycin (see page 4.5) and has grown well enough overnight to prepare a suspension, open the AN MicroPlate foil pouch and expose it to air for 20 minutes before inoculation. 2. For all species, keep the air exposure time consistent. Do NOT exceed 5 minutes between the time you begin and end inoculating AN MicroPlates for a single run. You should be able to inoculate six MicroPlates in 5 minutes. If the run requires more than six MicroPlates, use two people or divide the number of suspensions into smaller batches. 3. Wait 10 minutes after inoculating the AN MicroPlate before placing it in an anaerobic jar. This is necessary to slightly oxidize the buffer. • Rectangular jars can hold six or 24 MicroPlates, are easy to work with, and are inexpensive • 3.5 liter jars hold up to nine MicroPlates • 10.5 liter jars hold up to 28 MicroPlates and can stand upright 4. Use an anaerobic, atmosphere-generating system that does not produce hydrogen. Organisms with strong hydrogenases will reduce the tetrazolium in all wells if hydrogen is present. Add an anaerobic indicator strip to monitor the atmosphere. Call Biolog Technical Service for suggestions on where to find anaerobic supplies. 5. Incubate most anaerobes at 35° C for 20-24 hours. 6. After incubation, check the color of the MicroPlates before you open the anaerobic jar. MicroPlates should be clear with purple wells. Once you expose them to air, negative wells will slowly take on a faint green-blue color. If this green-blue color is present before you open the jar, the atmosphere was probably not fully anaerobic. Section 4 � Page 16 MicroStation System/MicroLog User Guide Apr 07
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DATABASE WINDOW Data Management and
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Data Management and Compiling Datab
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Arrow indicates selected entry Data
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Export Help Mark this choice if you
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8. Administration and Security In t
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Important! • If you test Password
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Log-In Privileges Administration an
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Remember that the password must be
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9. Technical Notes In this section:
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Making Biolog Universal Growth Agar
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� � � � Technical Notes Gen
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10. Frequently Asked Questions In t
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Q A Q A Q A Frequently Asked Questi
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Troubleshooting Symptom Cause Solut
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Troubleshooting Preparing Inocula S
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Troubleshooting MicroStation Reader
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Glossary Dendrogram Cluster diagram
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Glossary Spore, fungal Sterile, fun
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Culture Media Atmosphere Temperatur
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Appendices: Notes Notes: Section 13
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Appendix 4: Database Species Lists
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Appendix 5: Program Printouts Appen
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Appendix 5: Program Printouts 1�
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Appendix 5: Program Printouts Funga
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Appendix 5: Program Printouts Print
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Appendix 6: Dangerous Pathogens (DP
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