Reference no: EM133034390

Instructions

Your task:
To produce (1) a Methods section and (2) a Results section of a scientific paper. These will be based on the work undertaken on Lab 4: Bradford Protein Assay.

The PROTOCOL that you should re-write as Methods text is on pages 7 and 8. RESULTS TEXT exemplar. Included in this document, starting on page 9.

Instructions in the APPENDIX will lead you through the process of attaching a line of best fit to your standard curves, using Excel. This starts on page 11.

Composing the Methods section [see the protocol, pages 7 and 8]

Under the heading MATERIALS AND METHODS, describe what you did - in the past tense, using the passive voice, and without using personal pronouns (I, we).

You will need TWO sub-sections within the Methods: [1] Bradford protein assay; [2] Data Analysis.

These sub-headings already appear in the Homework 2 template Word file.

All of the general advice and guidance supplied in the Homework 1 Assignment Brief remains relevant here, so refer to that and to feedback received on that assignment. Consult PSB Reference Notes: Writing Scientific Papers, as required, for reminders of the key attributes of Methods text in a scientific paper.

Bradford protein assay
Be concise: no more than 200 words (there are about 365 words on this page) for this sub-section.
Here, you will "translate" the protocol, as seen starting on page 7, into a narrative
suitable for inclusion in the Methods section of a scientific paper. Some additional advice, specific to this task:
You prepared two sets of standards, each set using a different protein. You should write ONE, consolidated description of how these were prepared, making clear that there were two sets and identifying the proteins used. In so doing, describe the final composition without reporting the µL volumes of protein and water that were combined.
In the assay cuvettes, you combined a defined volume of sample (protein standard or test sample) with a defined volume of Bradford reagent. The sample volumes were invariant in all the assays. You should state this volume as well as the volume of Bradford reagent that was added to each cuvette.

Data analysis
Be concise: no more than 100 words for this sub-section.
Here, briefly describe how you made use of your standard curves to obtain estimates of the concentrations of protein in the test samples. If you obtained line equations for the linear parts of each standard curve (see Appendix, starting p. 11), these should be reported here. Do not provide "sample calculations".

Remember the essential attribute of an effective Methods section: it should permit a competent worker to REPEAT your study EXACTLY. This is the basis on which any Methods section will be judged.

Preparing the Results figure [see the data set, page 9]

In a section of your document headed RESULTS, provide a single figure, computer- drawn, containing standard curves-one based on BSA, the other on BGG-on a single set of axes. Include the full range of protein concentrations on this graph. Ensure that the data symbols for each standard protein are distinctive (distinguishing by colour alone is not sufficient), and are identified in a legend on the figure. Thin straight lines should connect the symbols to provide a visual indication of the trend for each plotted data set. DO NOT attach lines of best fit to this figure.

The figure should be laid out and annotated according to the principles of figure construction enumerated in PSB Reference Notes: Writing Scientific Papers. Make use of feedback received on Homework 1, as appropriate. Recall the basic guidance on graphing using Excel, found in the Homework 1 Assignment Brief, and guidance given in screencasts from earlier in the module.
Within your paper, place the figure after the Results text (see next)-if possible, on the same page (if not, on the page following).
Do not include ANY additional figures or tables in the Results section.

Composing the Results text

In the section of your document headed RESULTS, immediately under that heading, compose your Results text. This text should not exceed 300 words (a generous limit).

Three areas require attention within the Results text.
First, the text should describe, but not interpret or explain, the contents of the Figure. You must not just "read back" the data that are displayed on the Figure, but should draw to the reader's attention the most important and relevant details-particularly regarding the issue of the usable ranges of each standard curve. Remember: the usable range is that over which the response to increasing concentration of standard is linear. It is likely that you will NOT observe a linear response over the entire concentration range. Note also that it is not necessarily expected that the linear ranges for BSA and BGG will be identical-they may not be.

Second, you also must report in the text your estimate of the concentrations of protein in the two test samples. You should attempt to determine their respective protein concentrations against EACH of the two standards: that is, what do you get for X1 if determined via the BSA standard curve? Via BGG? Same for X2. If it is not possible to make a determination: you need to say so (and of course in such a case, you cannot give a numerical estimate!).

Third, related to the above, you should comment quantitatively on the accuracy of the reckonings of concentration for the two test samples.

The actual concentrations of the two test samples will have been revealed at (or just after) the Lab Session. You must indicate how well the estimated concentrations (as found via the assays) of X1 and X2 compared against the true concentration values for these solutions (again, provided it is possible to obtain estimates for these).
Remember: this is not a place to say anything about why one thing or another happened. Just make a factual comparison here.

Do not describe the LAYOUT of the graph: layout is obvious, Focus on observations emerging from the DATA.

The key variable (independent variable) to examine is µg/mL protein. What effect did increases in µg/mL of protein have on the dependent variable? Was this effect constant over the entire range of protein concentrations tested? Was it constant over part of the range? If the latter, over what range? Was the magnitude of the effect the same for both standard proteins? Are the slopes of the trendlines the same?
You will need to compare your X1 and X2 determinations (estimates) from each standard curve to the known true value of concentration in each. Of course, ideally, your "answer" from the Bradford assay should be very close to the true value, although you should not expect a "perfect" match.

An estimate that is within ± 5% of the true value can be considered "not different" from the true value.

If an estimate is indeed different from the true value: does the assay result represent an over-estimate or under-estimate? By how much?
Think about this in respect of how to express the degree of variance from the true value: Would it be better to report this as an absolute measure (e.g. difference) or a relative measure? (e.g. fraction or percentage of the true value)

Question: Record your cell counts for sample Loading Areas A and B in the table below. A systematic approach to counting is suggested by the (arbitrary) numbering of each 0.2 mm x 0.2 mm square.

Once you have counted all 10 squares, separately sum the values from sample Area A and from Sample Area B. Then, Sum the sums to obtain a grand total count (10 squares).

Square (0.2 mm x 0.2 mm) Smaple Area A Smaple Area B
1 (upper left)

2 (upper right)

3 (lower right)

4 (lower left)

5 (center)

Given that the grand total was obtained from 10 squares, Calculate and record below the average cell count (per square).

Referring the dimensions given in Activity 5.2.1 of the session Guide, Calculate the volume in cubic millimeters represented by each square.

Recognising that 1 mL = 1 cm3 = 1 cm x 1 cm x 1 cm, and that 1 cm = 10 mm, show workings bewlo to obtain the equivalent volume (1 ml) as cubic millimeters (mm3).

You should consider the above to be the "true values" against which you should compare the accuracy of the estimates obtained by your Bradford assay. Of note: both X1 and X2 were solutions of BGG.

Attachment:- Methods section.rar