1. Protocol title
The Title should indicate the goal of the protocol, providing enough details to help a prospective
researcher determine whether it is of interest. Keep in mind that your protocol could be useful for
researchers from a completely different field. When using specific terminology, make sure that the
title
can be understood by a broad range of scientists.
2. Author information
Provide complete affiliation information (department, research institution, city, country) for each
author.
3. Abstract
The Abstract should provide a brief summary of the protocol and highlight its goal and advantages. It
should also contain one or two sentences describing the context in which the assay/method was
developed.
4. Graphical overview
Consider including a graphical overview. It could be a flowchart of the method/procedures or a
graphical highlight of key steps, but it should not be a simple copy of other images in the
manuscript. See an
example here
from an article published in the
Bio-protocol journal.
5. Background
Briefly introduce the research area(s) that your protocol can be used to advance, providing the
rationale for developing the protocol. Place the protocol in the context of existing technology of the
research field, discussing previously used related methodologies, and summarizing the advantages,
limitations, and applications of your approach.
6. Materials and reagents
This section should provide a detailed list of all biological materials, reagents, solutions (with
recipes), and laboratory supplies required for the successful execution of the protocol, including
manufacturer catalog numbers, instructions on how to acquire/produce items that are not commercially
available, and storage conditions/shelf-life for critical reagents. Be precise about the ingredients
used, as a lack of clarity on minor details might lead to the failure of the experiment. Authors
should indicate whether any materials are subject to Material Transfer Agreements (MTAs) or if any
data are subject to Data Use Agreements (DUAs). Organize the items into four subcategories when
applicable: Biological Materials, Reagents, Solutions, and Recipes.
7. Equipment
List the equipment used in the experiments, including specific manufacturers and catalog/model
numbers.
8. Software and datasets
List individual software or datasets used in the experiments, including version and release date.
Provide references for items that are not commercially available.
9. Procedure
List all steps of the procedure chronologically, as it makes the protocol easier to follow. Be
specific in your instructions. Include details for material and reagents used (e.g., volume added,
specific container/tube, and incubation time) and the conditions in which the step is performed (e.g.,
temperature, agitation speed, and equipment settings); avoid using vague terms such as "several" or
"enough."
At crucial steps, include labels ("Caution," "Pause point," or "Critical," as appropriate) and notes
(see General Notes and Troubleshooting below), especially if it would increase the likelihood of
successfully repeating the experiment. As needed, include images and videos to clarify key steps and
representative data to illustrate the type of results obtained.
10. Data analysis
Provide a detailed description of data processing and analyses, including statistical tests, criteria
for data inclusion/exclusion, and details on the recommended number of biological and technical
replicates in each experiment. Highlight any specific skills necessary to perform the analyses (e.g.,
expertise using Linux, R, or other specific data analysis software).
When a detailed description of the data analysis method already appears in a published or posted
research article, summarize the analysis method and cite the publication or preprint. Clearly
indicate where the description of the analysis can be found (e.g., Supplemental information or
Figure X).
11. Validation of protocol
Provide evidence that the protocol is robust and reproducible. For experimental protocols, include
information about the number of replicates and controls that were used to validate the protocol.
Types of evidence:
- Reference specific data in a research article published
or posted by you or others (e.g., specific figures or supplementary materials in the article or
preprint).
- Provide data directly in this section of your preprint.
12. General notes and troubleshooting
Include comments that apply broadly to the protocol and are not fundamental to completing individual
steps. For example, address limitations of the protocol, its applicability to other experimental
systems or model organisms, or variability.
If stating common problems that might occur, describe ways to address them.
13. Acknowledgments
Acknowledge funding sources that have supported the work. If the protocol was adapted or modified from
previous work, acknowledge the relevant publications as well.
14. Competing interests
Provide a statement of financial and non-financial competing interests on behalf of all authors. For
more information, see the
Bio-protocol journal guidelines for disclaiming competing interests
here and a detailed list of
examples
at
PLOS.
15. Ethical considerations
All protocols that have used human and/or animal subjects should mention the specific ethics committee
that approved the experiment. Protocols including human subjects should also indicate that informed
consent was obtained from all subjects. Protocols including clinical trials should clearly state the
name of the trial registry and the clinical trial registration number in the manuscript. For animal
research, we recommend following the guidelines established
in Animal Research: Reporting of In Vivo
Experiments (ARRIVE). For more information, see the
Bio-protocol journal
publishing ethics notes.
16. References
Cite journal articles that are accepted, published, or posted as preprints. For in
press accepted articles, add the expected date of publication. Add the relevant citations for
all datasets, program code, and other resources used in the protocol.
17. Supplementary information
Include source or extended data and any relevant supporting materials in this section.
