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CAT #: 23090050

BCL2/JH Tube A - Unlabeled

Introduction

Master mixes are components of complete assays. Master mixes are composed of a buffered magnesium solution, deoxynucleotides, and multiple primers that target the gene segments of interest. Multiple primers are used to ensure a more comprehensive testing approach necessary to reliably identify clonal rearrangements. These tests are complete with the exception of Taq DNA Polymerase, which is not provided. A single thermocycler program and similar detection methods are used within each series of test to improve consistency, reduce human error, and facilitate cross training.

Standard Protocol:
50µl Total PCR Reaction Volume:

  1. Using gloved hands, remove the master mixes from the freezer. Allow the tubes to thaw; then gently vortex to mix.
  2. In a containment hood or dead air box remove an appropriate aliquot to clean, sterile microfuge tube (one tube for each of the master mixes). Aliquot volumes should be 45µl for each sample + 135µl for the positive, negative and no template controls. We recommend adding an additional 20µl to correct for pipetting errors.
  3. Add the appropriate amount of either AmpliTaq Gold or AmpliTaq DNA polymerase (0.25µl of either AmpliTaq Gold or AmpliTaq @ 5U/µl per 50µl total PCR reaction volume) to each of the master mixes and gently mix by inverting several times or gently vortexing.
  4. Aliquot 45µl of master mix to individual wells of a PCR plate.
  5. Add 5µl of DNA from the unknown and control samples to individual tubes or wells containing the respective master mix reactions, and pipette up and down several times to mix. Amplify target DNA using the universal thermocycler program.

Product Use

Historically, BCL2/JH t(14;18) Translocation Assays have been used to:

  1. Distinguish lymphoma from benign lymphoid hyperplasia
  2. Distinguish follicular lymphoma from other B-cell lymphomas that may have a similar appearance
  3. Monitor and evaluate disease recurrence and detect residual disease

Product Details

  • Specimen Requirements

    This product tests genomic DNA. DNA can be extracted from the following specimens:

    1. 5cc of peripheral blood, bone marrow biopsy, or bone marrow aspirate anti-coagulated with heparin or EDTA; OR,
    2. Minimum 5mm cube of tissue shipped frozen, at room temperature, or on ice in RPMI 1640; OR,
    3. Formalin-fixed paraffin embedded tissue or slides.
  • Description

    The BCL2 t(14;18) (q32;q21) translocation is found in 80-90% of follicular lymphomas and 30% of diffuse large cell lymphomas. The translocation is rarely present in other lymphoproliferative diseases. The t(14;18) brings about juxtaposition of BCL2 with the Ig heavy chain joining segment. This leads to a marked increase in expression of BCL2 driven by the Ig heavy chain gene enhancer. The bcl-2 protein inhibits programmed cell death (apoptosis) leading to cell accumulation. The majority of breakpoints on 18q21-22 occur within the major breakpoint region (Mbr) in the 3′ untranslated region of exon 3 (60-70% of the cases), and the minor cluster (mcr) region located 3′ to BCL2 exon 3 (20-25% of the cases). Some breakpoints occur at distant loci and will not be identified by this particular test. Therefore, a negative result does not completely exclude the presence of a BCL2/IGH gene rearrangement in the sample. In comparison, Southern blot analysis requires 1-2 weeks, is significantly less sensitive, and has more restrictive specimen requirements. The performance characteristics of this product have been independently determined by the EuroClonality Group.

    Four master mixes are included in this assay. Three are used to identify translocations in the major breakpoint region (Mbr) and minor cluster region (mcr) of BCL2. The Specimen Control Size Ladder master mix targets multiple genes and generates a series of amplicons of 100, 200, 300, 400, and 600 basepairs to ensure that the quality and quantity of input DNA is adequate to yield a valid result. This assay includes negative control DNA and positive control DNAs for both the Mbr and mcr. Clonality is indicated if any one of the master mixes generates clonal band(s). PCR products can be analyzed using standard gel electrophoresis with ethidium staining.

Disclaimer

©2019 Invivoscribe, Inc. All rights reserved. The trademarks mentioned herein are the property of Invivoscribe, Inc and/or its affiliates, or (as to the trademarks of others used herein) their respective owners.

For Research Use Only. Not for use in diagnostic procedures.

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