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Two-Dimension Electrophoresis Gallery

This page shows two-dimension electrophoregrams of various tissues from various laboratory animals, obtained in our laboratory.

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Methods in Two Dimension Electrophoresis

Sample preparation

Tissues/cells were suspended in protein extraction buffer (ReadyPrep™ protein extraction kit (Total Protein); Bio-rad), and homogenized with a bead-mill (TyssueLyser; Qiagen). After centrifugation, the supernatant was collected and its protein content was quantified with EZQ® Protein Quantification Kit (Invitrogen). The samples were reduced and alkylated with ReadyPrep™ reduction-alkylation kit(Bio-rad), and then applied for two-dimension electrophoresis.

First dimension: Isoelectic focusing

Samples were diluted with Rehydration/sample buffer (included in ReadyPrep™ 2-D starter kit; Bio-Rad), and used for isoelectric focusings (IEFs) with ReadyStrip™ IPG strips (pH3-10; Bio-rad) and PROTEAN® IEF cell(Bio-rad).

Second dimension: SDS-PAGE

After IEFs, the strips were treated with equilibrium buffers I and II (included in ReadyPrep™ 2-D starter kit; Bio-Rad), and then applied for SDS-PAGE. Precision Plus protein™ standard (unstained; Bio-rad) was used as molecular weight markers.

For SDS-PAGE, we used two gel systems:

  • 10-20% gradient gel (ReadyGel J; Bio-rad) + Tris Glycine SDS buffer (Bio-rad)
  • 4-12% gradient gel (NuPAGE® Novex BisTris Gel; Invitrogen) + MES buffer (Invitrogen)

Capture of gel images

Gels were stained with Sypro® Ruby gel stain (Invitrogen) and its images were captured with a laser scanner (FX-Pro; Bio-rad). The original images were converted into low-resolution jpegs with image software (Photoshop, Adobe) for easy handling on the web.

Gallery of 2DE images

Testis

AKR mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of testis from AKR mouse

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of testis from C57BL/6J mouse

DBA/1 mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of testis from DBA/1 mouse

Sperm

Epididymal sperm from C57BL/6Cr mouse

Sample: from an adult male (directly put into Rehydration/sample buffer)

2DE of epididymal sperm from C57BL/6Cr mouse

Ovary

Ovary from C57BL/6Cr mouse

Sample: from an 4-week-old female (stored in RNAlater ®, after buffer exchange using Zeba™ desalt spin column)

2DE of ovary from C57BL/6Cr mouse

Ovary from C57BL/6Cr mouse (48 h after PMSG treatment)

Sample: from an 4-week-old female (stored in Allprotect Tissue reagent, Qiagen).

Protein was extracted using AllPrep DNA/RNA/Protein Mini Kit (Qiagen) and used without reduction-alkylation treatment before IEF.

2DE of ovary from C57BL/6Cr mouse 48 h after PMSG treatment

Ovary from A/J mouse

Sample: from an 4-week-old female (stored in RNAlater ®, after buffer exchange using Zeba™ desalt spin column)

2DE of ovary from A/J mouse

Ovary from DBA/1 mouse

Sample: from an 4-week-old female (stored in RNAlater ®, after buffer exchange using Zeba™ desalt spin column)

2DE of ovary from DBA/1 mouse

Ovary from DBA/2 mouse

Sample: from an 4-week-old female (stored in RNAlater ®, after buffer exchange using Zeba™ desalt spin column)

2DE of ovary from DBA/2 mouse

Ovary from DBA/2 mouse (48 h after PMSG treatment)

Sample: from an 4-week-old female (stored in Allprotect Tissue reagent, Qiagen).

Protein was extracted using AllPrep DNA/RNA/Protein Mini Kit (Qiagen) and used without reduction-alkylation treatment before IEF.

2DE of ovary from DBA/2 mouse 48 h after PMSG treatment

Ovary from C3H/He mouse

Sample: from an 4-week-old female (stored in RNAlater ®, after buffer exchange using Zeba™ desalt spin column)

2DE of ovary from C3H/He mouse

Ovary from BDF1 mouse

Sample: from an 4-week-old female (stored in RNAlater ®, after buffer exchange using Zeba™ desalt spin column)

2DE of ovary from BDF1 mouse

Ovary from BDF1 mouse (48 h after PMSG treatment)

Sample: from an 4-week-old female (stored in Allprotect Tissue reagent, Qiagen).

Protein was extracted using AllPrep DNA/RNA/Protein Mini Kit (Qiagen) and used without reduction-alkylation treatment before IEF.

2DE of ovary from BDF1 mouse 48 h after PMSG treatment

Ovary from SD rat

Sample: from a non-pregnant adult female (stored in RNAlater ®; Its estrous stage was not monitored.)

2DE of ovary from SD rat

Oocytes

Ovulated oocytes from BDF1 mice

Sample:Ovulated oocytes from BDF1 mice treated with eCG and hCG (directly put into Rehydration/sample buffer).

2DE of ovulated oocytes from BDF1 mice

In vitro matured oocytes from BDF1 mice

Sample:In vitro matured oocytes from 21-day-old BDF1 mice (directly put into Rehydration/sample buffer).

2DE of in vitro matured oocytes from BDF1 mice

In vitro matured oocytes from C57BL/6Cr mice

Sample:In vitro matured oocytes from 21-day-old C57BL/6Cr mice (directly put into Rehydration/sample buffer).

Molecular Standard: Mark12™ Wide Range Unstained Standard (Invitrogen).

2DE of in vitro matured oocytes from C57BL/6Cr mice

In vitro matured oocytes from DBA/2 mice

Sample:In vitro matured oocytes from 21-day-old DBA/2 mice (directly put into Rehydration/sample buffer).

2DE of in vitro matured oocytes from DBA/2 mice

In vitro matured oocytes from ICR mice

Sample:In vitro matured oocytes from 21-day-old ICR mice (directly put into Rehydration/sample buffer).

2DE of in vitro matured oocytes from ICR mice

Liver

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of liver from C57BL/6J mouse

AKR mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of liver from AKR mouse

Heart

C57BL/6Cr mouse

Sample: from an adult male (stored at -80°C)

2DE of heart from C57BL/6Cr mouse

JW rabbit (Ventricle)

Sample: frozen tissue purchased from Funakoshi Co., Ltd.

2DE of heart from JW rabbit

Hartley guinea pig (Ventricle)

Sample: frozen tissue purchased from Funakoshi Co., Ltd.

2DE of heart from Hartley guinea pig

SD rat (Ventricle)

Sample: frozen tissue purchased from Funakoshi Co., Ltd.

2DE of heart from SD rat

Syrian Hamster (Ventricle)

Sample: from an adult female (stored in RNAlater ®)

2DE of heart from Syrian Hamster

Skeletal muscle

C57BL/6Cr mouse, thigh muscle

Sample: from an adult male (stored at -80°C)

Molecular Standard: ECL Dual-vue Western Blotting Marker (GE Healthcare).

2DE of skeletal muscle from C57BL/6Cr mouse

Kidney

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of kidney from C57BL/6J mouse

Urine

BDF1 mouse

Sample: from an immature female, without reduction-alkylation treatment before IEF

2DE of urine from BDF1 mouse

Lung

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of lung from C57BL/6J mouse

Spleen

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of spleen from C57BL/6J mouse

Thymus

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of thymus from C57BL/6J mouse

Cerebrum

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of cerebrum from C57BL/6J mouse

Cerebellum

C57BL/6J mouse

Sample: from an adult male (stored in RNAlater ®)

2DE of cerebellum from C57BL/6J mouse