Products

Bullet Semi-Dry Transfer One

Nacalai Tesque has developed a transfer buffer for use with western blotting that allows for fast semi-dry protein transfer.
* This product is suitable for use with our Bullet PAGE One precast gel. It may not work as expected with other products.

Features

  • Protein transfer in 10 min
  • High transfer efficiency with a wide range of molecular weights
  • No gel equilibration necessary
  • Ready to use

>> Information on Bullet PAGE One Precast Gel

>> Bullet Blocking One for Western Blotting

Product Information

Performance Comparison

Bullet Semi-Dry Transfer One was compared to the conventional methods by Kyhse-Andersen (1984) and Towbin (1979). Under the same conditions, it achieved higher transfer efficiency.

Transfer buffer  Bullet Semi-Dry Transfer One Kyhse-Andersen Towbin
Conditions 10V (constant), 10 min
Membrane
image

*1 Kyhse-Andersen J.1984. J. Bio chem. Biophys. Methods 10:203-209
Composition: 48 mM Tris, 39 mM Glycine, 20% Methanol, 0.04% SDS
*2 Towbin H., Staehelin T., and Gordon J. 1979. Proc. Natl. Acad Sci 76:4350-4354.
Composition: 25 mM Tris, 192 mM Glycine, 20% Methanol

Conditions:

Electrophoresis gel Bullet PAGE One Precast Gel 5-15%（#13079）
Samples
  1. Protein Ladder One (#09547), 5 µl added to each lane
  2. Protein Markers (10x) (#29458), 2 µl added to each lane
  3. Protein Markers (10x) (#29458), 3x diluted, 2 µl added to each lane
  4. Protein Markers (10x) (#29458), 10x diluted, 2 µl added to each lane
  5. Protein Markers (10x) (#29458), 30x diluted, 2 µl added to each lane
Membrane PVDF
Transfer conditions 10V (constant voltage), 10 min
Detection CBB Stain One (#04543)

Applications

Detection by antigen-antibody reaction

β-Actin's strong signal yielded a clear band using only 10V for 10 min to transfer proteins. A weaker signal, such as COX4's, may be detected more easily by increasing the voltage.

Protein β-Actin COX4
Voltage 10V 10V 25V
Transfer time 10 min
Chemiluminescence
image

Method

Sample

Proteins extracted from HL-60 cells using RIPA Buffer (#08714).

Lane 1 : 10 µg
Lane 2 : 3 µg
Lane 3 : 1 µg

Summary of procedures

The above results were obtained using our Bullet series to speed up electrophoresis and western blotting.

Protein Products used Time
Electrophoresis Bullet PAGE One Precast Gel 5-15%（#13079） 10 min
Semi-dry protein
transfer		 Bullet Semi-dry Transfer One（#15353）
*PVDF membrane		 10 min
Blocking Bullet Blocking One（#13779) 5 min
Primary antibody
reaction		 Anti β-Actin(C4)(Mouse), Monoclona（l Santa Cruz #sc-47778） diluted 1000x 60 min
Anti-COX4(D-20)(Goat)(Santa Cruz #sc-69359） diluted 1000x
Secondary antibody
reaction		 Anti-Mouse IgG(Goat), HRP Conjugate（Santa Cruz #sc-2005） diluted 10,000x 60 min
Anti-Goat IgG(Bovine), HRP Conjugate（Santa Cruz #sc-2350） diluted 10,000x
Chemiluminescence
detection		 Chemi-Lumi One Super（#02230) 5 min. exposure

Protocols

How To Use

Materials required

Western blotting filter paper (ex: Whatman® 3MM Chr , 70mm×85mm) 12 sheets
Transfer membrane (ex: PVDF membrane, 70mm×85mm) 1 sheet

Pre-wetting the membrane ]

  1. Soak the membrane with methanol (or ethanol) in tray.
  2. Remove methanol (or ethanol) from the tray, add 100 mL of water and shake.

Transfer

Note: Use 40-50 mL of transfer buffer for one Bullet PAGE One Precast Gel.
Carefully assemble the gel, membrane and filter papers to avoid bubbles.
  1. Soak 6 sheets of filter paper in transfer buffer and place on anode plate of a blotting apparatus.
  2. Place membrane on filter papers.
    Note: Remove excess water with a paper towel.
  3. Wash the gel with water for 10 seconds (within 20 seconds) and place on the membrane.
    Note: Remove excess water with a paper towel.
  4. Soak 6 sheets of filter paper in transfer buffer and place on the gel.
  5. Set the blotting apparatus and connect to a power supply.
  6. Transfer proteins from the gel to the membrane with constant voltage (10-25V) for 10 minutes (refer to the below table).

Transfer condition for one Bullet PAGE One Precast Gel

Constant voltage Time Maximum current
25V 10 min approx. 1.2A
10V 10 min approx. 330mA

In case of insufficient transfer at 10V for 10 minutes, transfer at 25V (10 minutes), or extend the transfer time.

Downloads

Bullet Semi-Dry Transfer Onepdf (PDF 2.6 MB)

Ordering Information

Product Name Storage Product No. PKG Size Price
Bullet Semi-Dry Transfer One Room Temp. 15353-01 1 L e-Nacalai

Related Products

Bullet PAGE One Precast Gel

Product Name Wells Storage Product No. PKG Size Price
Bullet PAGE One Precast Gel, 5-11% 13 4-8 °C 13077-04 10 Sheets e-Nacalai
Bullet PAGE One Precast Gel, 5-11% 17 4-8 °C 13078-94 10 Sheets
Bullet PAGE One Precast Gel, 5-15% 13 4-8 °C 13079-84 10 Sheets
Bullet PAGE One Precast Gel, 5-15% 17 4-8 °C 13080-44 10 Sheets
Bullet PAGE One Precast Gel, 6% 13 4-8 °C 13081-34 10 Sheets
Bullet PAGE One Precast Gel, 6% 17 4-8 °C 13082-24 10 Sheets
Bullet PAGE One Precast Gel, 8% 13 4-8 °C 13083-14 10 Sheets
Bullet PAGE One Precast Gel, 8% 17 4-8 °C 13084-04 10 Sheets

Bullet Blocking One for Western Blotting

Product Name Storage Product No. PKG Size Price
Bullet Blocking One for Western Blotting 4-8 °C 13779-14 200 ml e-Nacalai
13779-01 1 L

(Storage) RT: Room temperature, A: Cool and dark, R: Refrigerator, F: Freeze