Erratum: Rapid Analysis of Chromosome Aberrations in Mouse B Lymphocytes by PNA-FISH

DOI:

10.3791/5771

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Summary

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A correction was made to: Rapid Analysis of Chromosome Aberrations in Mouse B Lymphocytes by PNA-FISH.

Abstract

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A correction was made to: Rapid Analysis of Chromosome Aberrations in Mouse B Lymphocytes by PNA-FISH. Table 1 (the list of solutions to prepare) was omitted from the Protocol section. The contents of Table 1 are as follows:

SolutionReagentsCommentsStorage
B Cell Wash Buffer500 ml 1x Hanks Balanced Salt SolutionFilter through 0.22 μm stericup filtration unit4 °C
5 ml 50 °C heat inactivated FCS
5 ml 100x Pen/Strep
B Cell Medium418.5 ml RPMI-1640Filter through 0.22 μm stericup filtration unit4 °C
50 ml 50 °C heat inactivated FCS
5 ml 100x Pen/Strep
5 ml glutamine
5 ml nonessential amino acids
5 ml sodium pyruvate
1.8 ml 14.2 M 2-mercaptoethanol
5 ml 1M HEPES
Fixative3:1 v/v methanol/glacial acetic acidMake fresh
0.075 M KCl1.395 g KClRoom Temperature
250 ml distilled H2O
FISH Master Mix40 ml 50% Dextran sulfateVortex the solution and leave in a shaking platform overnightAliquot and store at -20 °C
20 ml 20x SSC
40 ml sterile distilled H2O
70% Formamide/2x SSC10 ml 20x SSCAdjust to pH 7 with 1M HClAliquot and store at -20 °C
20 ml distilled H2O
70 ml deionized formamide
0.01 M HCl1 ml 1M HClAdjust to pH 2.0Room Temperature
99 ml distilled H2O
1x PBS/MgCl250 ml 1M MgCl2Room Temperature
950 ml 1x PBS
50% Formamide/2x SSC20 ml 20x SSCAdjust to pH 7.25
Formamide used in this step does not have to be deionized.
Make fresh
80 ml distilled H2O
100 ml formamide
4x SSC/0.1% Tween2050 ml 20x SSCRoom Temperature
200 ml distilled H2O
250 μl Tween20
DAPI staining solution (80 ng/ml)40 μl 0.2mg/ml DAPI stock4 °C in light protected staining jar
100 ml 2x SSC
Mowiol Mounting Medium6 g glycerolIncubate at 50 °C with stirring for 2hrsAliquot and store at 4 °C
2.4 g Mowiol 4-88
6 ml distilled H2O
12 ml 0.2 M TrisCl pH 8.5
230 μl 1% Thimerosal (w/v in water)

Table 1. List of solutions. 

Protocol

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$$\rightleftharpoonup{xx}$$ $$\longleftharp{xx}$$, $$\longrightharp{xx}$$,

A correction was made to: Rapid Analysis of Chromosome Aberrations in Mouse B Lymphocytes by PNA-FISH. Table 1 (the list of solutions to prepare) was omitted from the Protocol section. The contents of Table 1 are as follows:

SolutionReagentsCommentsStorage
B Cell Wash Buffer500 ml 1x Hanks Balanced Salt SolutionFilter through 0.22 μm stericup filtration unit4 °C
5 ml 50 °C heat inactivated FCS
5 ml 100x Pen/Strep
B Cell Medium418.5 ml RPMI-1640Filter through 0.22 μm stericup filtration unit4 °C
50 ml 50 °C heat inactivated FCS
5 ml 100x Pen/Strep
5 ml glutamine
5 ml nonessential amino acids
5 ml sodium pyruvate
1.8 ml 14.2 M 2-mercaptoethanol
5 ml 1M HEPES
Fixative3:1 v/v methanol/glacial acetic acidMake fresh
0.075 M KCl1.395 g KClRoom Temperature
250 ml distilled H2O
FISH Master Mix40 ml 50% Dextran sulfateVortex the solution and leave in a shaking platform overnightAliquot and store at -20 °C
20 ml 20x SSC
40 ml sterile distilled H2O
70% Formamide/2x SSC10 ml 20x SSCAdjust to pH 7 with 1M HClAliquot and store at -20 °C
20 ml distilled H2O
70 ml deionized formamide
0.01 M HCl1 ml 1M HClAdjust to pH 2.0Room Temperature
99 ml distilled H2O
1x PBS/MgCl250 ml 1M MgCl2Room Temperature
950 ml 1x PBS
50% Formamide/2x SSC20 ml 20x SSCAdjust to pH 7.25
Formamide used in this step does not have to be deionized.
Make fresh
80 ml distilled H2O
100 ml formamide
4x SSC/0.1% Tween2050 ml 20x SSCRoom Temperature
200 ml distilled H2O
250 μl Tween20
DAPI staining solution (80 ng/ml)40 μl 0.2mg/ml DAPI stock4 °C in light protected staining jar
100 ml 2x SSC
Mowiol Mounting Medium6 g glycerolIncubate at 50 °C with stirring for 2hrsAliquot and store at 4 °C
2.4 g Mowiol 4-88
6 ml distilled H2O
12 ml 0.2 M TrisCl pH 8.5
230 μl 1% Thimerosal (w/v in water)

Table 1. List of solutions.

Disclosures

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$$\rightleftharpoonup{xx}$$ $$\longleftharp{xx}$$, $$\longrightharp{xx}$$,
No conflicts of interest declared.

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Tags

PNA FISH AnalysisSolution PreparationB Cell Wash BufferB Cell MediumFixative Solution0 075 M KClFISH Master Mix70 Formamide 2x SSC