Total protein lysates were mixed with sample buffer and heated at 95 °C for 5 min. Equal amounts of protein were subjected to SDS/PAGE and transferred to nitrocellulose. The membranes were blocked in protein‐free PBS blocking buffer (Thermo Scientific) for 1 h at room temperature. Primary and secondary Ab incubations were performed in protein‐free blocking buffer/0.05% Tween‐20 for 1 h at room temperature. For all membranes, protein signals were detected using an Odyssey imaging instrument and analyzed using instrument software (Li‐Cor Biosciences, Lincoln, NE, USA).
Pbs blocking buffer
Manufactured by Thermo Fisher Scientific
Sourced in United States
PBS blocking buffer is a solution used to reduce non-specific binding in various immunoassay and molecular biology applications. It contains a mixture of proteins and other components that help block undesired interactions between the target analyte and the assay surface or other components. This product is commonly used to prepare samples and control backgrounds in techniques such as Western blotting, ELISA, and immunohistochemistry.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using pbs blocking buffer
1
Protein Separation and Detection
2
SARS-CoV-2 Spike Protein ELISA Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Purified SARS-CoV-2 full length spike or RBD protein was diluted to 1 μg/mL in PBS. Maxisorp plates (Nunc) were coated with 100 μL per well (100 ng protein per well) and incubated overnight at 4°C. Plates were washed 3x with PBST (0.1% Tween) and blocked with 100 μL casein in PBS blocking buffer (ThermoFisher) for 1 hour at room temp. Plates were again washed 3x with PBST (0.1% Tween), and 100 μL of serum samples, serially diluted 2 fold in casein in PBS blocking buffer, in duplicate, was added to the wells and incubated at room temperature for 1 hour. Plates were washed 4x with PBST (0.1% Tween), and 100 μL secondary antibody, rabbit anti-human IgG Fc HRP (Novus Biologicals, NBP1-73529) diluted 1:4000 in casein in PBS blocking buffer, was added to the wells and incubated for 1 hour at room temperature. The wells were washed 5x with PBST (0.1% Tween) and developed with the KPL TMP 2-component peroxidase substrate kit (Seracare, 5120-0047). The reaction was stopped with KPL stop solution (Seracare, 5150-0020) and read at 450 nm. The threshold for positivity was calculated as the average plus 3 times the standard deviation of negative control sera. Reported titers are the reciprocal value of the highest dilution at which signal was observed above the calculated threshold.
3
Western Blot Protein Detection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Equal amounts of total protein lysates were mixed with sample buffer, heated at 95 °C for 5 min, and loaded to SDS/PAGE and transferred to nitrocellulose. The membranes were blocked in protein‐free PBS blocking buffer (Thermo Scientific, Rockford, IL, USA) for 1 h at room temperature. Primary antibodies were incubated in the blocking buffer for 16 h at 4 °C. Secondary antibodies were incubated in the same blocking buffer for 1 h at room temperature. Protein signals were detected using an Odyssey imaging instrument, and the intensities of the immunoreactive bands were quantified by densitometric analysis using instrument software (Li‐COR Biosciences, Lincoln, NE, USA).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!