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Immunocytochemistry is used to validate the antibody staining and for assessing and validating the protein expression pattern in selected human cell lines.
A Western blot analysis is performed on a panel of human tissues and cell lines to evaluate antibody specificity. For antibodies with uncertain result a revalidation using an over-expression lysate is performed.
Immunocytochemistry is used to validate the antibody staining and for assessing and validating the protein expression pattern in selected human cell lines.
For each antibody, the observed staining in the different cell lines is assigned a validation score based on concordance with available experimental gene/protein characterization data in the UniProtKB/Swiss-Prot database. The validation scores for up to three cell lines are merged into one of the main categories; Supported, Approved, or Uncertain to represent the overall antibody staining in all analyzed cell lines.
Immunofluorescent staining of human cell line U-2 OS shows localization to vesicles.
Immunofluorescent staining of human cell line MCF7 shows localization to the Golgi apparatus.
Antibody dilution
1:25
1:53
Literature conformityi
The immunostaining patterns is compared for consistency with available experimental gene/protein characterization data in the UniProtKB/Swiss-Prot database and other experimental evidence for location described in scientific literature.
The subcellular location is partly supported by literature or no literature is available.
The subcellular location is partly supported by literature or no literature is available.
WESTERN BLOTi
A Western blot analysis is performed on a panel of human tissues and cell lines to evaluate antibody specificity. For antibodies with unreliable result a revalidation using an over-expression lysate is performed.
Western Blot is used for quality control of the polyclonal antibodies generated in the project. After purification, the antibodies are used to detect bands in a setup of lysate and different tissues. The result is then scored Supported, Approved, or Uncertain.
Band of predicted size in kDa (+/-20%) with additional bands present.
Uncertain
Analysis performed using a standard panel of samples. No bands detected.
Figure description
Lane 1: Marker [kDa] 250, 130, 95, 72, 55, 36, 28, 17, 10 Lane 2: Negative control (vector only transfected HEK293T lysate) Lane 3: Over-expression Lysate (Co-expressed with a C-terminal myc-DDK tag (~3.1 kDa) in mammalian HEK293T cells, LY414362)
Lane 1: Marker [kDa] 230, 130, 95, 72, 56, 36, 28, 17, 11 Lane 2: RT4 Lane 3: U-251 MG Lane 4: Human Plasma Lane 5: Liver Lane 6: Tonsil
Target mass (kDa)
67.8, 67.4
67.8, 67.4, 58.4, 46.6, 42.6, 17.4
Antibody dilution
1:250
1:250
PROTEIN ARRAYi
A protein microarray containing 384 different antigens including the antibody target is used for analysis of antibody specificity.
A protein array containing 384 different antigens including the antibody target is used to analyse antibody specificity. Depending on the array interaction profile the antibody is scored as Supported, Approved, or Uncertain.
