Palladium colloid
For research use only. Not for clinical diagnosis.
Cat. No. WRC-WRPD1-40NM-EX
Palladium Colloid
BACKGROUND
Immuno-chromatography test
Left: palladium colloid
Right: gold colloid
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Test line on the left specimen can be seen more clearly under the same condition.
Easy conjugation
Degrees of conjugation of antibodies to metal colloids depend on the palladium or gold colloids applied.
Lower incidence of agglutination due to higher negative charge than conventional metal colloids
High concentration: more than 12 in OD525
Can be preserved for as long as six months in a refrigerator (4 - 10ºC)
Feature
The palladium colloid is highly concentrated (OD525 ＝ 12) compared with other
High concentration
commercially available gold products (OD525＝ 1). Therefore, efficient preparation is
Stable
possible using small instruments, even on a large scale
The product may be stored for six months at room temperature (15～25°C).
Large antibody binding
The colloids can hold up to 1.5 times as many antibodies as other commercially
capacity
available gold colloids.
Optimal pH may be achieved during the colloid-antibody reaction simply by the addition
Easy pH adjustment
of buffer solution.
Shelf Life
Six months after purchase date
4℃
Storage Method
Instructions
prior
to
A bottle is shaken before use.
use
Note
We recommend that small scale tests be conducted and evaluated before scaling up to
production.
Product List
Catalog No.
WRC-WRPD1-40NM-EX
Application
Product Name
Palladium Colloid for Immunochromatography, 40 nm
Volume
10 ml
50 ml
100 ml
Preparation of an antibody-noble metal conjugate using new gold colloid
or new palladium colloid
www.cosmobio.co.jp
Palladium colloid
Materials
- Our gold colloid solution, either undiluted or diluted (×2 or ×5) with 10 mM Tris pH 9.2
(henceforth denoted Tris) (620 µL/experiment), is used for preparation of the antibody
conjugate. In order to determine the optimal conditions, Tris pH 7.0 and pH 8.0 were
also used in the preparation. (5 mM Tris or 2 mM borax may be sufficient instead of 10
mM Tris.)
- Tris (100 µL) is added to 100 µL of gold colloid solution.
Note: For the antibody solution, a 0.1~0.2 mg/mL solution diluted with Tris, or a solution
of 30-60 µg/mL, is used. (Because of the strength of the antigen–antibody reaction, a
solution of 30–60 µg/mL is used for expensive antibodies.) For example: antibody
solution; Candida albicans (rabbit), Cosmo Bio, VIS, product No. 6411.
To prepare a 25 µg/mL antibody solution (120 µL) from an antibody solution of 1.5
mg/mL, 120 µL of Tris is added to 1.8 µL of the antibody solution.
- To prepare a 1% BSA solution (5 mL), 50 mg of BSA is added to 5 mL Tris (600µ L/
experiment.
- To prepare a 1% PEG solution, 9.9 g of water is added to 100 mg PEG (molecular
weight 20,000) (50µL/experiment) - To prepare a 1% BSA–PEG mixed solution, 1%
PEG solution and 1% BSA solution are mixed at a ratio of 9:1 (400µL/experiment)
- To prepare a 2.5% sodium caseinate solution (1 mL), 1 mL Tris is added to 25 mg of
sodium caseinate.
- To prepare 1 mL of gold colloid stabilizing solution (5% D (+)-trehalose•2H2O solution
(100µL/experiment)), 1mL of BSA solution is added to 50 mg of 1% trehalose 2-hydrate
(100µL/ experiment).
Method
1. An undiluted gold colloid solution (or a 2× dilution with Tris) and an antibody solution
are mixed at a ratio of 1:1. For example, 100 µL of gold colloid solution and 100 µL of
antibody solution are mixed in a 0.5 mL tube. The use of a siliconized tube is
recommended
Notes: When the gold colloid solution is mixed with an antibody solution containing a
salt, it may become blue or purple. The antibody may be centrifuged using a centrifugal
filter (Microcon, Millipore) in order to remove the salt. For antibody solutions not
containing salts, the gold colloid solution is mixed with the antibody solution in small
quantities (100 µL) to begin with.
2. The mixture is allowed to stand for 15 minutes, and then 200 µL of BSA–PEG mixed
solution is added. PEG prevents the conjugate from sticking to the tube. Instead of l%
BSA solution, 2.5% sodium caseinate solution (10 mM Tris pH 10.0) may be sufficient.
3. The mixture is centrifuged for 15 minutes using a force of 15,000 × g for WRGH1 (50
nm), 5,000 × g for WRGH1 (60 nm), 15,000 × g for WRGH2, or 30,000 × g for WRGM3.
Since the bottom will come off a 0.5 mL tube at 15,000 × g, the 0.5 mL tube is put inside
a 1.5 mL tube.
4. After removal of the supernatant, BSA–PEG mixed solution (200 µL) is added. If the
mixture sticks to the tube and is not distributed evenly, the number of rotations should
be lowered. The supernatant is again removed.
5. BSA–PEG solution (200 µL) is added again and the mixture is centrifuged to remove
the supernatant.
For research use only. Not for clinical diagnosis.
Distributor
Manufacturer: Winered Chemical Corporation
TOYO 2CHOME, KOTO-KU, TOKYO, 135-0016, JAPAN
URL: http://www.cosmobio.co.jp e-mail: [email protected]
[Outside Japan]
Phone : +81-3-5632-9617
FAX : +81-3-5632-9618
[国内連絡先]
Phone : +81-3-5632-9610
FAX : +81-3-5632-9619
Palladium colloid
6. The gold colloid stabilizing solution (5% D (+)-trehalose 2-hydrate solution) (100 µL)
is added and mixed.
Note: The antibody concentration and the pH of the buffer solution vary depending on
the antibody used. Although room temperature preservation may be sufficient for the
gold colloid, other reagents and solutions should be stored in the refrigerator.
Immunochromatography kit
Materials
1. The gold–antibody conjugate is applied to a fiberglass conjugate pad. For example,
when using a Millipore GFCP203000 pad, 10 µL of the conjugate is applied at the tip,
and the apparatus is dried at room temperature. Example of absorption pad: Millipore
CFCS203000
2. Swabbing of antibody on membrane
- Membrane: Hi-Flow Plus HF 180, Millipore.
- IgG dilution solution: Casein (10 mg) is added to 5 mL of 10 mM phosphoric acid
buffer (pH 7.0), and the mixture is warmed to cause dissolution. The mixture is then
filtered using a 0.45 µm filter and diluted × 4 with water.
- IgG (primary antibody): antibody (e, g.1.46 mg/mL) is usually available in 1 mL portion
(refer to catalog). To prepare 15 µL of a 0.1 mg/mL solution, 1 µL of IgG is added to 13.6
µL of IgG dilution solution.
- Anti-rabbit IgG antibody (secondary antibody): 1 mg/mL (VECTOR), usually available
in 1 mL portions (refer to catalog). To prepare 15 µL of a 0.1 mg/mL solution, add 1.5 µL
of anti-IgG to 13.5 µL of IgG dilution solution.
- 0.5% Blocking solution: casein (150 mg) is added to 20 mM phosphoric acid buffer (pH
7.8) (30 mL) and the mixture is warmed to cause dissolution. The mixture is then filtered
using a 0.45 µm filter.
- 3% Sucrose solution: sucrose (0.9 g) is added to distilled water (30 mL).
Method
1. IgG is applied to a membrane (4x40mm), 1 mm in width, as shown in the figure.
2. The membrane is dried for 30 to 60 minutes at room temperature.
3. The membrane is dipped in the blocking solution and gently shaken for 20 minutes.
4. The membrane is then dipped in distilled water and shaken. (Repeat.)
5. The membrane is then dipped in the sucrose solution and shaken gently for 10
minutes.
6. Any excessive moisture on the membrane surface is sucked up with paper, and the
membrane is dried at room
temperature.
Application of IgG and
secondary antibody to a
membrane: The apparatus is a
combination of a sample pad, a
conjugate pad, a membrane, and
an absorption pad on a backing
sheet (NIPPN. Techno Cluster,
Inc.).
For research use only. Not for clinical diagnosis.
Distributor
Manufacturer: Winered Chemical Corporation
TOYO 2CHOME, KOTO-KU, TOKYO, 135-0016, JAPAN
URL: http://www.cosmobio.co.jp e-mail: [email protected]
[Outside Japan]
Phone : +81-3-5632-9617
FAX : +81-3-5632-9618
[国内連絡先]
Phone : +81-3-5632-9610
FAX : +81-3-5632-9619