1. How critical are the incubation times for the Crystal panels?
The results could be affected if the panels are not incubated for the proper times as follows:
ENTERIC/NONFERM 18-20 hours GRAM POSITIVE 18-24 hours RAPID GRAM POSITIVE 4 hours ANAEROBE 4 hours NEISSERIA/HAEMOPHILUS 4 hours
Note: Panels should be read within 30 minutes of removing from the incubator and they should be held at room temperature. Longer or shorter incubation times can result in false reactions.
2. What other tests are required for Crystal panels?
ENTERIC/NONFERM OXIDASE, INDOLE GRAM POSITIVE GRAM STAIN RAPID GRAM POSITIVE GRAM STAIN ANAEROBE GRAM STAIN, CATALASE, INDOLE NEISSERIA/HAEMOPHILUS GRAM STAIN
Note: The use of BBL DMACA Indole is strongly suggested. Weakly positive organisms may yield false negative results with other formulations such as Kovacs' reagent. Catalase testing should be performed using 15% H2O2 with 1% Tween 80. Use of 3% H2O2 for anaerobic organisms can yield false negative results.
3. What types of media can be used with Crystal ID Systems?
ENTERIC/NONFERM TSA W/5% SB, MAC GRAM POSITIVE TSA W/5% SB, COLUMBIA W/ 5% SB, PEA W/5% SB, COLUMBIA CNA W/5% SB RAPID GRAM POSITIVE TSA W/5% SB, COLUMBIA W/5% SB, PEA W/5% SB, COLUMBIA CNA W/5% SB ANAEROBE CDC ANAEROBE BLOOD AGAR, BRUCELLA BLOOD AGAR, COLUMBIA BLOOD AGAR, SCHAEDLER BLOOD AGAR, TSA W/5% SB NEISSERIA/HAEMOPHILUS CHOCOLATE, TSA W/5% SB, COLUMBIA, NUTRIENT AGAR, MARTIN LEWIS, MOD. THAYER MARTIN, NEW YORK CITY, V AGAR, GC LECT™ Agar
Note: All must be pure cultures 18 to 24 hours old. If a customer cannot use one of the media listed in our insert, our recommendation is to "qualify" their media for use with Crystal by testing the quality control battery in the insert and an additional twenty organisms. Based on these test results, the user can decide whether the medium can be used with the Crystal system. However, the performance of the product cannot be guaranteed if other media are used.
4. How do I avoid bubbles in the wells?
A humid environment is essential for optimal performance. If the account is having trouble with bubbles forming, then check the humidity conditions of the incubator and correct if necessary. We recommend a humidity level of 40 to 60%. Make sure a pan of water is in the bottom of the incubator. Other recommendations include:
The above measure is only needed for cases where the humidity conditions are insufficient and bubbles are a problem. In most cases, bubbles do not interfere with color-change interpretations.
- Place the tray on the shelf directly above the filled water pan in the incubator.
5. I am not getting an ID. What should be done?
This usually occurs because mixed cultures are utilized. We recommend that purity plates be inoculated from the panel inoculum suspension during the evaluation process. This may also occur if improper inoculum concentrations are used; check inoculum concentration.
6. Are Crystal™ Systems AOAC approved?
Not yet, but in the near future we anticipate receiving approval for Enteric/Nonfermenter and overnight Gram Positive.
7. Can the inoculated broth sit for several hours before inoculating the panels?
No. We recommend that the broth be utilized promptly after adjustment to the appropriate McFarland standard.
8. Which McFarland standards can be used to adjust the inoculum for Crystal?
ENTERIC/NONFERM <0.5? GRAM POSITIVE 0.5 RAPID GRAM POSITIVE 2 ANAEROBE 4-5 NEISSERIA/HAEMOPHILUS 3
*One well isolated large (2-3 mm or larger in diameter) colony (or 4-5 smaller colonies of the same morphology).
Vortex inoculum broth well to break up any clumps (this creates a homogeneous suspension). Use only the inoculum broth provided or saline to adjust the McFarland if it is too high. Do not use water or nutritive broth. Remove the excess amount added to the tube with a sterile pipette so that the final volume of inoculum fluid is approximately equivalent to the original volume in the tube.
9.Is the Crystal™ Lightbox U.L. (Underwriter Laboratories) approved?
Yes.
10. Once the foil pouch is opened, the package insert states that the panel must be used promptly. Define "promptly".
The panel must be used within 1 hour after opening. The reagents on the prongs may accumulate moisture and start to degrade after 1 hour without the desiccant and false reactions may result.
11. For the Crystal panels that use fluorescent reactions, can I read them with a Wood's lamp?
The panel reader uses the same wavelength as a Wood's lamp (365 nm); however, because the reaction wells are small, distance from the light sources and the angle of the light can be critical when reading Crystal ID panels. For this reason, we recommend using the Panel Viewer.
12. When comparing databases for each of the media, taxa are missing from some of the databases. For example, in the Crystal Anaerobe the organisms Bacteroides gracilis, Clostridium putrificum and Fusobacterium gonidiaformans are in the Schaedler database but not in the CDC database. Why?
The reason for the slight differences in taxa between databases is due to the variability of growth and reactivity of organisms on the different media.
13. What can I use to suspend the organism in the inoculum broth?
Use an inoculating loop or sterile cotton swab. Do not use polyester swabs as they may yield false results. If swabs are to be used, be sure to squeeze all broth from the swab before inoculating the panel.
14. What is the proper environment for incubation of the panels?
The panels must be incubated aerobically without CO2 in a humidity- controlled environment at 35-37ºC. Incubation with added CO2 or anaerobically can yield false results. Lack of humidity can cause bubbles in the wells that may make reading difficult. Furthermore, the panels should be incubated upside down (the way they are read) to provide the optimum mix of broth and substrate. Trays should not be stacked more than two high.
It is also important that the incubator door remain closed during incubation. The door can be opened briefly for a maximum of three times, if necessary. Repeated opening and closing may cause temperature fluctuations, resulting in false reactions.
15. How do I interpret the fluorescent reactions?
For fluorescent reactions, only wells which are brighter than 4A should be considered positive. Well 4A is a fluorescent negative control; any reactions equal to or less than (not as bright as) 4A should be considered negative.
CRYSTALSPEC™ NEPHELOMETER
1. What range of turbidity does the CrystalSpec™ Nephelometer read?
The CrystalSpec™ Nephelometer was designed to measure the turbidity of microbial suspensions equivalent to McFarland Standards 0.5 through 4.0.
2. Do I need to purchase the calibration standards separately?
No. The Calibration Blank and CrystalSpec™ Calibration Standard is included with the nephelometer. A separate catalog number exists (245015) in the event the standards are lost, broken or they have expired.
3. Can I use any standards to calibrate my instrument?
No. The meter must be calibrated with the supplied CrystalSpec™ Calibration Blank and CrystalSpec™ Calibration Standard. The use of other standards will result in miscalibration of the instrument.
4. In what kinds of broths and tubes can I make my suspensions?
Organisms may be suspended in water, saline or clear growth medium of yellow to pale brown color (e.g., Mueller Hinton Broth, TSB or BHI Broth). Use of other colored liquids as a suspending medium is not recommended. Clear flat bottom glass tubes 16 - 17 mm diameter must be used for samples.
5. What sample volume should I use?
If the sample volume is between 2.0 mL and 4.0 mL insert the Low Volume Sample Adapter. If the volume is greater than 4.0 mL, then the Low Volume Sample Adapter is not needed. Volumes below 2.0 ml cannot be measured.
6. Can I use the standards as a visual read?
No.
7. What kind of warranty does the nephelometer have?
The warranty period is one year from the date of purchase. Defective product will only be replaced at no charge during the warranty period.
8. How often do I calibrate?
The meter should be tested with the calibration standard on a daily basis. If the calibration standard reads 2.0 0.1, the instrument is calibrated and does not need recalibration. The calibration should be stable if ambient conditions of temperature and light remain relatively constant. If these conditions change, recalibration is recommended. Units must be recalibrated after replacing the battery.
9. What maintenance is required?
The only maintenance that is required is to change the battery when a "B0" or "E" is displayed, indicating a low battery (one 9V alkaline battery is included).
10. What does an "E" mean on the display?
If the "E" is displayed the internal error checking has detected a problem due to improper use, discharged battery or an electronic failure. If calibration and/or replacement of the battery does not correct the problem, contact BDDS Product Support.
11. What will happen during the calibration procedure if I lift my finger off the calibration button before pressing the test button?
You will get a reading of "0.0 McFarland". The calibration process should be repeated.
12. What will happen if I try to read a specimen above a 4.0 McFarland?
The instrument will display an "E".
13. What is the minimum dating guaranteed for the standards?
Six months.
14. Does BD have a repair program for the nephelometer?
No. Defective instruments will replaced with a new instrument only during the warranty period. Broken instruments are not refurbished.
