Western blotting is an important technique used in cell and molecular biology. By using a western blot, researchers are able to identify specific proteins from a complex mixture of proteins extracted from cells.In this technique a mixture of proteins is separated based on molecular weight, and thus by type, through gel electrophoresis. These results are then transferred to a membrane producing a band for each protein. The membrane is then incubated with labels antibodies specific to the protein of interest.

The unbound antibody is washed off leaving only the bound antibody to the protein of interest. The bound antibodies are then detected by developing the film. As the antibodies only bind to the protein of interest, only one band should be visible. The thickness of the band corresponds to the amount of protein present; thus doing a standard can indicate the amount of protein present.

Tanda offers professional Western blotting services tailored for proteomics Western blot applications for biotech and pharma companies. Our service covers sample processing, blotting, quantitative imaging, data analysis and professional reporting.

Western Blot Service Workflow
Total Turnaround Time: 1-2 Weeks

Sample collection and shipping
Western blotting's success depend on sample quality. Keep the samples fresh and avoiding protein degradation is necessary. It is also important to provide enough samples so there are ample proteins to load for each lane. Please contact us for the collection, handling and shipping procedures of common sample types including plasma, serum, whole blood, tissue, cells, lysates and urine, as well as shipping address, shipping time and a Service Form.

Sample processing
We offer full service for WB and can handle sample processing for you. If needed, we can perform protein extraction/lysis, sample filtration (which removes cell debris and is important for reducing non-specific reactions), and BCA. You can also do your own sample processing and send us lysates.

Phase 1: antibody Optimization
Commercial antibodies are not always validated for every sample type. We usually start our project with primary antibody dilution optimization. By default we will test 3 concentrations of the primary antibody on positive samples. Please indicate on your sample manifest which samples should be used for optimization.

If you prefer, we can also work with a dilution ratio that you have already optimized and skip this phase.

Main experiments
The main experiment(s) will run all your samples. If you have a particular experiment layout, please include it in your inquiry and we will follow it. By default we use 15 lane gels and load 10ug of total protein each lane. Therefore we can process up to 14 samples per gel and we charge by the number of gels run.

Data analysis
After the main experiments are done, we will generate a professional report (see sample report at the end of page) which contains the experiment data as well as the conditions of the experiment, and information about reagents and instruments used. Our Western blots can be quantitative at no additional cost.

Service options explained
Quantification: We can normalize your target of interest's signal against either a loading control antibody or a fluorescent total protein stain to give quantitative result for your WB.

Total protein stain: in short, we need to use this option if we need to load more than 10ug of total proteins per lane, due to target protein expression being low in the samples. Total protein stain is a better alternative to using loading control antibodies for quantification. The main drawbacks for using loading control are that first the signal's linear range is too short and saturates easily, that if we load more than 10ug of total protein per lane it usually results in saturated loading control antibody signal and renders the data useless. Also your treatment to cells may affect your loading control gene's expression level, contrary to common assumption that loading control gene expression states constant, and more and more journals are starting to require evidence showing that the treatment does not affect loading control gene expressions. Total protein stain, on the other hand, has excellent linear range and is a more stable way to normalize in quantitative Western blotting.

Antibody optimization: we will conduct a preliminary study using 3 different concentrations of your antibodies on positive samples to decide the best concentration to use for later experiments. The positive samples to use in optimization needs to be indicated in the sample manifest as "use for optimization". This is usually a necessary phase and only if the customer can provide exact dilution ratios to use can we skip this phase.

Return or dispose excess materials: The remaining samples and reagents be shipped back to you or disposed of. There are fees associated with both options.

Sample type and processing requirements: different sample types require different processing. Details see below.

*Clarification: filtering the sample to get rid of cellular debris. Helps with reducing WB background.
**Serum and Plasma: 1-1.5uL of sample per lane, protein concentration assumed to be ~50mg/mL, need clarification if samples are lipemic
***Blood: need to spin down to serum/plasma, extra charge
****Low expression protein: Need to concentrate protein. Extra charge.

Sample replication: by default we assume all samples are tested in singlet.

Sample role in experiment: the 3 most common sample roles in WB are:
1. used in optimization
2. used as positive or negative control
3. a regular sample, aka a Unknown

Special requirements: Contact us for anything not covered in the Service Form.

High Sensitivity and Specificity: With high-quality antibodies and optimized experimental procedures, it can specifically detect low-abundance target proteins, effectively avoiding non-specific signal interference even in complex samples, ensuring the accuracy and reliability of detection results.

Comprehensive Detection Capability: It can not only detect conventional protein expression levels but also analyze post-translational modifications such as phosphorylation, glycosylation, and methylation. Additionally, it is suitable for various biological samples and different types of proteins, providing customers with comprehensive protein analysis services.

Strict Quality Control: Every step, from sample processing to result analysis, strictly follows standardized operating procedures and a quality control system to ensure the reproducibility and stability of experimental results. Furthermore, the company has an internal quality inspection mechanism that conducts batch sampling and validation for each set of experimental results.

Customed Service Solutions: Based on customers' specific research needs and sample characteristics, customized experimental solutions are provided, including antibody selection and experimental condition optimization, ensuring that the results meet research requirements to the greatest extent.

Professional Technical Team: The company has an experienced and highly skilled technical team with strong backgrounds in molecular biology and immunology. All team members are well-versed in every aspect of WB (Western Blot) detection technology, offering full-process technical support and consulting services, from experimental design, sample processing, and operation to result analysis and interpretation.