WesternBlots -
ImmunoBlots
General Information about WesternBlot Kits
MATERIALS PROVIDED
|
Test unit |
8 Tests |
|
Nitrocellulose-strips loaded with nuclear and mitochondrial antigens |
8 strips |
|
Blocking- buffer (cloudy), containing PBS, blocking protein and 0,1 % acide, ready for use |
1 vial - 20 ml |
|
Goat-anti-human-IgG-Conjugate (conjugated with alkaline phosphatase), Concentrated |
1 vial - 0,1 ml
|
|
Substrate (BCIP), ready for use |
1 vial - 10 ml |
|
Stop solution (3 % acetic acid), ready for use |
1 vial - 20 ml |
|
Serum dilution and washbuffer (SDW-buffer), containing PBS, proteins, Tween 20 and 0,1 % acide, 10x Concentrated |
1 vial - 20 ml |
TECHNICAL DATA
|
Specimen: |
Serum |
|
Required sample size: |
20 µl per test |
|
Incubation time: |
approx. 1,5 hours at room temperature |
|
Storage: |
at 2 to 8 °C (refrigerator) |
|
Test unit: |
8 strips |
Technical notes and qc-criteria
Do not change anything of the recommended protocol! Temperatures below 18 °C and above 28 °C may affect the development of the bands. In the case of a too high temperature, the substrate incubation should be watched with care and stopped sooner than in the recommended protocol to avoid an overdyeing of the bands. The opposite applies in the case of too low temperatures. The recommended time for substrate incubation is a guideline only.
Store all kit components at 2 – 8 °C in a refrigerator!
The components supplied in this kit are intended for use as an integral unit. Therefore, the components of different lots should not be mixed. This is especially true for the reference (template) delivered in the kit. This reference (template) can be used for the strips of the kit only.
Do not use kits or components after the expiry date printed on the outer box.
Efficient rinsing of uncomplexed serum components is a fundamental requirement of the immunoblot procedures. Therefore, the washing procedure should be performed particularly carefully. Insufficient washing will cause aberrant results and high background staining.
QC-criteria
These immunoblots are manufactured in lots of 10 (20) strips. Two of these strips are developed with control sera. One of the developed strips is delivered in each kit. The other strip is documented at the manufacturer´s. Using these two marker strips, the reference (template) is made. The reference (template) indicates the exact positions of the antigens on the undeveloped strips. Therefore, the running of controls is not necessary to analyse the band pattern.
To control the proper performance of each strip, a control band is dotted 1 cm below the upper end. This band must develop in any case. If this band does not develop, or only very faintly, one of the reagents was not added or has deteriorated.