Citations & References

Quantitative quality control in microarray image processing and data acquisition.

  • Authors: Wang X, Ghosh S, Guo SW
  • Journal: Nucleic Acids Res 2001; (29):15 E75-E75
  • PubMed ID: 11470890

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Brochure: Quanti-Cult Brochure

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Citations & References

Evaluation of quantitative miRNA expression platforms in the microRNA quality control (miRQC) study.

  • Authors: Mestdagh P, Hartmann N, Baeriswyl L, Andreasen D, Bernard N, Chen C, Cheo D, D'Andrade P, DeMayo M, Dennis L, Derveaux S, Feng Y, Fulmer-Smentek S, Gerstmayer B, Gouffon J, Grimley C, Lader E, Lee KY, Luo S, Mouritzen P, Narayanan A, Patel S, Peiffer S, Rüberg S, Schroth G, Schuster D, Shaffer JM, Shelton EJ, Silveria S, Ulmanella U, Veeramachaneni V, Staedtler F, Peters T, Guettouche T, Wong L, Vandesompele J
  • Journal: Nat Methods 2014; (11):8 809-815
  • PubMed ID: 24973947

Citations & References

Comparison and verification of quantitative competitive reverse transcription polymerase chain reaction (QC-RT-PCR) and real time RT-PCR for avian leukosis virus subgroup J.

  • Authors: Kim Y; Gharaibeh SM; Stedman NL; Brown TP
  • Journal: Journal of Virological Methods 2002 2-Jan:1-8
Catalog #
  • 4318739(Discontinued)
  • 4327058(Discontinued)
  • 4327059(Discontinued)

Product FAQ

How do you prepare blood cells for chromosome analysis?

Answer

Phytohemagglutinin Assay:

Lymphocytes are differentiated cells which normally do not undergo subsequent cell divisions. By culturing lymphocytes in the presence of a mitogen (KaryoMAX Phytohemagglutinin (M-Form) (PHA), Cat. No. 10576), they are stimulated to replicate their DNA and enter into mitosis. After an optimum time of the cells being cultured (46 h for a newborn and 68 h for an adult), a mitotic inhibitor, KaryoMAX COLCEMID Solution (Cat. No. 15210 or 15212), is added to the lymphocyte culture for 20 min. The addition of COLCEMID to dividing cells acts to prevent the synthesis of spindle fibers and, therefore, to stop mitosis in metaphase. Metaphase is the optimum phase of mitosis for microscopically visualizing the chromosomes. By submitting cells to a hypotonic solution and a series of fixation steps, metaphase chromosomes can be microscopically observed and analyzed.

As a quality control measure, each lot of Phytohemagglutinin is tested as a chromosome reagent for the examination of metaphase spreads used for cytogenetic studies. This reagent is evaluated by supplementation to an approved, non-phytohemagglutinin containing chromosome medium. These samples are then supplemented with freshly collected human peripheral blood and have been found to be acceptable in their ability to produce blastogenesis with human lymphocytes when compared to a previously tested control.

Test Procedure:

1. The required volume of peripheral blood is collected aseptically in a sodium heparinized vacutainer tube or syringe.
2. Add 10 ml of either PB-MAX Karyotyping Medium (Cat. No. 10386) to each sterile T-25 flask to be set up for the assay.
3. Add 0.75ml blood to each tube.
4. Incubate flask in CO2 incubator for 48 to 68 h with caps loose.
5. Add 0.05-0.1 ug/ml COLCEMID to each flask for a 15 minute incubation.
6. After 15 min, transfer flask contents to a 15 ml centrifuge tube and spin down at 1,200 rpm for 5 min.
7. Remove supernatant and resuspend pellet.
8. Add 10 ml of 0.068 M KCl to pellet and gently mix. Allow to sit at room temperature for 15 min. Add 0.5 ml of fixative (three parts absolute methanol to one part glacial acetic acid). Gently mix with pipette.
9. Centrifuge for 5 min at 1,200 rpm. Aspirate off supernatant. Add 10 ml of fixative, mix, and let sit at room temperature for 10 min.
10. Repeat centrifugation step. Add 5 ml of fixative, mix, and let sit for 10 min at room temperature.
11. Centrifuge and aspirate off supernatant. Add 5 ml of fixative and incubate for 10 min at 4°C.
12. Centrifuge at 1,200 rpm for 7 min. Aspirate off supernatant. Gently resuspend in fixative.
13. Prepare slide by placing 6 to 8 drops of cell suspension on slide. Use warming plate to dry slides.
14. Once all traces of moisture have disappeared, you can begin staining procedure:
- Stain with Giemsa Stain for 6 to 8 min, or any other appropriate stain.
- Remove slides from stain and rinse under running distilled water.
- Allow slides to thoroughly dry and examine slides for well-spread metaphases.

Record number of cells and at the same time record the number of these cells which are in metaphase. At least 500 cells must be counted for a valid assay. Statistically determine the Mean Mitotic Index (a quantitative measure) and Mean Banding Resolution (a qualitative measure). The test sample values must favorably compare to the reference control values.

Answer Id: E4306

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Product FAQ

How do TaqMan Gene Expression Assays help save time?

Answer

The TaqMan Gene Expression Assays enable researchers to conduct real-time quantitative PCR gene expression studies quickly and easily by eliminating the time-consuming processes involved in assay development, which typically include the following stages:
- Sequence selection
- BLAST analysis against transcript and genomic databases
- Primer and probe design
- Oligonucleotide ordering/manufacture
- Optimization of primer and probes concentrations
- Quality control and functional testing

Answer Id: E1838

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Product FAQ

Which RNA isolation kit do you recommend using with Arcturus Laser Capture Microdissection (LCM) samples?

Answer

The Arcturus PicoPure RNA Isolation Kit (Cat. Nos. KIT0204, KIT0214) is specifically designed to consistently recover high-quality total RNA from fewer than 10 cells, even from a single cell. This kit is designed to provide consistent, efficient RNA recovery without sacrificing quality. Small volume elution allows you to maximize your recovery of RNA from small numbers of cells for use with your gene expression analysis experiments.

For LCM cells from paraffin sections, we recommend using the Arcturus Paradise Plus RNA Extraction and Isolation Kit (Cat. No. KIT0312I) that unlocks RNA from formalin-fixed, paraffin-embedded (FFPE) tissues. As little as 5 ng of the isolated fixed RNA can then be amplified using reagents and methods that have been specially optimized to overcome the challenges of cross-linked FFPE templates.

We also recommend the Arcturus Paradise PLUS QC Kit (Cat. Nos. KIT0303, KIT0313). This kit offers a simple solution for evaluating the quality of RNA from formalin-fixed paraffin-embedded (FFPE) tissues before proceeding with LCM and downstream molecular analysis. The reagents provided in the kit are optimized to enable efficient extraction, isolation, and reverse transcription of total RNA from FFPE tissue scrapes. The RNA quantity and quality is then measured using quantitative real-time PCR with a housekeeping gene like beta actin.

Answer Id: E15454

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