When you are writing a short report, you need to try to get out of the habit of writing up your experiment under the traditional style of aim, methods, results and discussion. This applies particularly to the methods stage which is NOT written as a separate section but is included in the LEGEND which accompanies the RESULTS.
The main problems students have with writing the legend are deciding on:
- what content or information to put in and how much to put in;
- how to structure this content;
- where to put this content.
Remember that there is particular emphasis on the detailing of the conditions under which the data were obtained. WE WANT CONDITIONS NOT PROCEDURES. Sufficient methodological information must be provided to enable the reader to see how the experiments were performed and to allow critics to repeat and/or pick holes in the experimental approach.
The methods in a short report are written as LEGENDS. Legends sit under your graphs and figures but above your tables. The information in a legend should tell the reader how you obtained the data in the figure/table by answering the following questions:
- What were the conditions or circumstances under which the data was obtained, e.g. the concentrations, pH, temperature etc during the reaction?
- How was the experiment performed?
Legends provide enough information to allow the reader to repeat the experiment in their lab and with their equipment.
This information tells the reader the conditions or circumstances under which the data was obtained; concentrations, pH, temperature, wave length etc.
Figures 1 and 2. Effect of high-and low-cholesterol diets on blood cholesterol concentration. Subjects (Table 1) consumed a high (n=3) or low (n=4) cholesterol diet for up to 3 months. Plasma obtained from samples obtained at the times shown were analysed for cholesterol by incubation in 100 mM Tris-HCl, pH 7.7 containing 1 mM aminoantipyrine (AAP), 6 mM phenol, 1 mM sodium cholate, 2.8 U/ml peroxidase, 7.4 mM Triton X-100, 0.65 U/ml cholesterol esterase and 0.45 U/ml cholesterol oxidase. After 10 min at room temperature, the absorbance of the incubation mixture was determined at 500 nm.
This information tells the reader in what way or how the experiment was performed and when or for how long certain actions were carried out.
The typical structure of the legend in a short report is shown in the following table.
|Stage 1||Explanation of how the data in the figure/table were obtained. You need to describe the conditions under which the data were obtained and, in general, how the experiment was performed.|
|Stage 2 (optional)||Summary of experimental results (but don’t repeat what is in the discussion or in the tables and/or figures).|
Figures 1 and 2. Effect of high- and low-cholesterol diets on blood cholesterol concentration.
Subjects (Table 1) consumed a high (n=3) or low (n=4) cholesterol diet for up to 3 months. Plasma obtained from samples obtained at the times shown was analysed for cholesterol by incubation in 100 mM Tris-HCl, pH 7.7 containing 1 mM aminoantipyrine (AAP), 6 mM phenol, 1 mM sodium cholate, 2.8 U/ml peroxidase, 7.4 mM Triton X-100, 0.65 U/ml cholesterol esterase and 0.45 U/ml cholesterol oxidase. After 10 min at room temperature, the absorbance of the incubation mixture was determined at 500 nm.
Stage 2: (Optional)
The extinction coefficient of the red colour produced when AAP is oxidised by the hydrogen peroxide produced in the cholesterol oxidase reaction, was 1 mM-1 cm-1. *P<0.01 for difference between low- and high- groups.
Changing Procedures into Legends
Procedures or protocols give you a step-by-step method for carrying out your experiment. Often they are lists of instructions for making up materials (solutions etc.) and what to do with these materials. When you are writing up your report, you need to change the protocol into a legend. This means you have to focus on the conditions under which the experiment was performed and not the detailed procedures you followed to make solutions etc. In this way you will give a more general description of your materials and methods.
Example Procedure: a method for measuring the activity of the enzyme alcohol dehydrogenase in liver.
- The assay has a total volume of 1.0 ml and contains 0.5 ml 0.8 M glycine, pH 9.5, 50 ml of 60 mM NAD, 50 ml of 1 M ethanol and distilled water to 980 ml.
- Place this mix in a 1ml cuvette and zero the spectrophotometer at 340 nm. 20 ml of a liver extract is then added. This extract was prepared by homogenising 400 mg of liver with 4 ml of a homogenising buffer, containing 50 mM Tris pH 7.5 and 1 mM dithiothreitol (DTT) to stabilise the enzymes.
- Then measure the absorbance at 340 nm every 15 s for 5 min.
- Convert the linear rate, expressed as a change in absorbance per min, to the number of umoles of product (NADH) per min using the extinction coefficient for NADH at 340 nm, 6.2 mM-1 cm-1.
How to Change a Procedure into a Legend
Step 1. You need to have a clear understanding of the concept behind the experiment by asking yourself the following question.
How does this experiment measure the activity of alcohol dehydrogenase?
The activity of alcohol dehydrogenase, an enzyme found in liver, was measured by the rate of appearance of NADH, monitored by an increase in absorbance at 340 nm using the e = 6.2 mM-1 cm-1.
Step 2. You need to think about what happened in the experiment so that you can order the information in a logical way in the legend. Ask yourself the following questions.
What reacted with what?
Alcohol dehydrogenase in liver reacted with NAD and ethanol.
What was the product?
NADH was the product.
What is the relationship between the product and the activity of alcohol dehydrogenase?
The rate of appearance of NADH monitored by an increase in absorbance at 340 nm using the e = 6.2 mM-1 cm-1 indicated the activity of alcohol dehydrogenase.
Step 3. You need to calculate the final concentration of each of the components of the assay, keeping the proportions constant. Ask yourself the following questions.
What is the final glycine concentration if 0.5 ml of 0.8 M glycine is used?
0.4 M glycine, pH 9.5 (the pH won’t alter significantly on dilution)
What are the final NAD and ethanol concentrations?
3 mM NAD and 50 mM ethanol
What is the final concentration (g/ml) of the liver homogenate?
0.1 g liver extract per 1 ml of buffer solution - the fact that the researcher used 400 mg doesn’t really matter as long as the proportions are kept constant.
What is the final concentration of the liver extract in the reaction mixture?
2 mg liver tissue per ml of reaction mixture.
Liver homogenates (0.1g wet weight/ml prepared in 50 mM Tris, pH 7.5, 1 mM DTT) were assayed in 0.4 M glycine pH 9.5, containing 3 mM NAD and 5 mM ethanol. The assay contained 2 mg liver tissue/ml reaction mixture. The progress of the reaction was monitored by the increase in absorbance at 340 nm over a 5 minute time period and the reaction rate calculated using the extinction coefficient for NADH at 340 nm, ε = 6.2 mM-1 cm-1.
You have now reached the end of the Legend section.