الوصف
Protein A/G-agarose
beads Price:
U.S.＄**0.***1ml; $**0—5ml; $***0—*5ml
(negotiable) CAT. No.: P***3
Storage conditions: 4℃, valid for one year.
Do not freeze. Product Description
Protein A+G Agarose is mainly used for
immunoprecipitation (IP) or co-immunoprecipitation (Co-IP), and can
also be used for antibody purification.
Protein A is a cell wall surface protein found
in Staphylococcus aureus with a molecular weight of *2kDa; Protein
G is an immunoglobulin binding protein expressed by C-type or
G-type streptococcal bacteria. Protein A and Protein G have similar
functions, and their binding abilities to different immunoglobulin
subclasses are different. Appropriately recombinant Protein A and G
are combined with agarose gel in a certain way, which can be used
for immunoprecipitation or antibody
purification. Protein A+G Agarose is suitable for
immunoprecipitation of all antibodies that can be
immunoprecipitated by Protein A Agarose and Protein G Agarose
alone, including human IgG1, IgG2, IgG3, IgG4, mouse IgG1, IgG2a,
IgG2b, IgG3, rat IgG1, IgG2a, IgG2b, IgG2c, rabbit IgG, rabbit,
goat polyclonal antibodies.
The Protein A and Protein G in this product are
covalently linked to 4% high cross-linking, high-flow agarose and
mixed in a 1:1 ratio. Each ml of Protein A+G agarose beads
(precipitate) can bind more than *0mg human IgG. The average
diameter of agarose beads in this product is *****5μm, the
recommended linear flow rate for antibody purification is
******0px/h, and the maximum pressure resistance index is
0.1MPa. If this Protein A+G Agarose is used for
conventional immunoprecipitation, each ml of this product can be
immunoprecipitated *0 times.
Notes 1. Do not freeze this
product. 2. Protein A+G Agarose must be fully
resuspended before use, that is, fully inverted several times to
mix evenly. 3. This product contains trace amounts of
preservatives, which will not affect conventional
immunoprecipitation and antibody purification. However, if enzyme
activity determination is involved later, it is advisable to wash
Protein A+G agarose beads three times with appropriate solutions
such as TBS before using this product to fully eliminate possible
interference caused by preservatives.
4. Starting from protein sample collection,
protein samples must be operated at 4 degrees or on ice in all
steps. 5. This product is limited to scientific
research by professionals, and shall not be used for clinical
diagnosis or treatment, shall not be used for food or medicine, and
shall not be stored in ordinary
residences. 6. For your safety and health, please wear a
lab coat and disposable gloves when
operating. Instructions:
1. Immunoprecipitation
(IP): a. Preparation of protein
samples: (a) For adherent cells in a *0 cm cell culture
dish, remove the cell culture medium, wash once with PBS, and then
add **0ul to 2ml of cell lysis buffer to lyse the cells. You can
use Western and IP cell lysis buffers produced by LABLEAD or
various RIPA lysis buffers to lyse the
cells. (b) For tissue samples, refer to the proportion
of lysis buffer used for adherent cells for
lysis. (c) For suspended cells, collect the cells by
centrifugation, wash once with PBS, and then refer to the lysis
method of adherent cells for lysis.
Note: For detailed lysis methods, refer to the
detailed usage of different lysis buffers. For different culture
equipment, refer to the amount of lysis buffer used for *0 cm
culture dishes for lysis. If the concentration of the protein
sample obtained by lysis is too high, it can be appropriately
diluted with lysis buffer or PBS. If the concentration of the
protein sample is too low, it is advisable to appropriately reduce
the amount of lysis buffer in the subsequent lysis process. b.
Removal of non-specific binding
(optional): (a) Take **0ul to 1ml protein sample, the
protein amount is about **0ug to 1mg, add about 1ug of common IgG
of the same species as the IgG used in immunoprecipitation and *0ul
of fully resuspended Protein A+G Agarose, and shake slowly at 4ºC
for *0min to 2h. (b) Centrifuge at ***0rpm (about ***0g) for
5min, and take the supernatant for subsequent
immunoprecipitation. Note: The so-called IgG of the same species
means that mouse IgG is used in the subsequent immunoprecipitation.
In this step, normal mouse IgG can be added. If normal IgG is not
available, other mouse IgG antibodies that do not affect subsequent
detection can be added. By incubating with normal IgG and Protein
A+G Agarose, non-specific binding can be fully reduced and
background can be reduced. c.
Immunoprecipitation: (a) Add 0.**2ug of the primary antibody used
for immunoprecipitation and shake slowly at 4ºC
overnight. (b) Add *0ul of fully resuspended Protein A+G
Agarose and shake slowly at 4ºC for **3h (to facilitate subsequent
washing operations, the amount of fully resuspended Protein A+G
Agarose can be adjusted to *0ul).
(c) Centrifuge at ***0rpm (about ***0g) for
5min, or centrifuge at high speed for a short time, and carefully
remove the supernatant. Note that it is better to leave a small
amount of supernatant than to remove Protein A+G
Agarose. (d) Wash the precipitate 5 times with the lysis
buffer or PBS used when preparing the protein sample, and the
amount of lysis buffer or PBS used each time is 0.**1ml. The
centrifugation conditions and supernatant removal requirements
during washing are the same as step
(c). (e) After the last wash, remove the
supernatant, add ****0ul 1X SDS-PAGE electrophoresis loading buffer
Vortex to resuspend the precipitate, and centrifuge at high speed
for a short time to centrifuge the sample to the bottom of the
tube. (f) Treat at **0ºC or in a boiling water bath
for **5 minutes, and take part or all of the sample for SDS-PAGE
electrophoresis. Temporarily unused samples can be stored at
**0ºC. 2.
Co-immunoprecipitation: Refer to the immunoprecipitation method, but
co-immunoprecipitation (Co-IP) usually must use fresh protein
samples that have not been frozen. Although ordinary
immunoprecipitation can use frozen protein samples, it is also
better to use fresh protein samples.
3. Antibody
purification: a. Preparation:
(a) Filter the solution used with a 0.*5um or
0.2um pore size filter membrane.
(b) All solutions must be degassed (degas) by
ultrasound or other methods.
(c) Select an appropriate purification column
and fill the purification column with an appropriate amount of
Protein A+G Agarose. You can also use LABLEAD's pre-packed column
products. (d) Wash and balance the purification column
with ****0 column volumes of TBS. The flow rate can be controlled
by a constant flow pump to 1 ml/min (1 ml pre-loaded column). If
there is no constant flow pump, the purification column can also be
washed and balanced entirely by
gravity. b. Antibody
purification: (a) Load the purification column with the
antibody to be purified. (b) After the antibody to be purified passes
through the column, wash with ****0 column volumes of TBS to remove
unbound and non-specifically bound proteins. Whether the washing is
complete can be determined by measuring the absorbance at **0
nm.
| بلد: |
China |
| نموذج رقم: |
-
|
| سعر فوب: |
(قابل للتفاوض)
أحصل على آخر سعر
|
| الموقع: |
China |
| سعر الحد الأدنى للطلب: |
- |
| الحد الأدني للطلب: |
- |
| تفاصيل التغليف: |
bottle |
| موعد التسليم: |
7-15 days if in stock |
| القدرة على التوريد: |
- |
| نوع الدفع: |
T/T, L/C, D/A, D/P, Western Union, Money Gram, PayPal |
| مجموعة المنتج : |
Protein Biology
|
