Biology Question

Science

ashford university

Question Description

I need someone to answer page 2 and 3 of the attached biology document............................

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BIO273 – Experiment 1 – Absorption Spectrum & Standard Curve for Biuret Assay of BSA. Step 1: Start with an 8 mg/mL Bovine Serum Albumin (BSA) solution, prepare a series of 5 standards that range in concentration from 0-8 mg/mL (do not include 0 or 8 mg/mL as any of your standards). Fill in the table on the assignment sheet and briefly summarize below how you will prepare each standard. An example is given. The total volume of each dilution will be 4 mL. You will make them directly into the cuvettes that go into the spectrophotometer. Note the diluent is a phosphate-buffered saline (PBS) solution, not water. Total volume Volume of stock Volume of PBS to Concentration (mg/mL) being prepared Dilution Factor needed (mL) be added (mL) (mL) 2* 4 1 3 1/4 4 4 4 4 8 (STOCK already made) *In this example, I want to make a 2 mg/mL dilution. 2 mg/mL is ¼ of the working stock concentration of 8 mg/mL (I did 2 divided by 8 to get 1/4). If I am doing a 1/4 dilution and making 4 mL total, then 4 x 1/4 = 1 mL. I need to pipette 1 mL of the 8 mg/mL solution into the test tube. To make 4 mL total, I need to add 3 mL of water to this. (Subtract 4 mL total - 1 mL stock = 3 mL). Repeat this for each of the concentrations you choose. The higher the concentration, the more stock you will use. If your values are too weird - like need 3.666666 mL, then choose a different concentration or dilution factor, since this is not practical and will lead to error. Try to keep the volumes to whole numbers or decimals that do not repeat. Please note the concentration unit throughout this entire experiment is always mg/mL (milligrams per millilitre). You do not need to do any unit conversions. We are always keeping this same unit for concentration. Summarize how you will prepare each of the standards below: Example: Standard 1: Concentration ____2 mg/mL______ Pipette 1 mL using 1000 µL micropipette into a cuvette labelled “2”. Pipette (new tip) 3 mL using 1000 µL micropipe into same cuvette, mix with that tip. Assignment Sheet – Standard Curve of Biuret Protein Assay for Quantification of BSA 1. Preparation of Standards [4 marks] Standard 1 Concentration (mg/mL) Total volume Volume of Volume of PBS being prepared stock needed to be added Dilution Factor (mL) (mL) (mL) 2* 2 3 4 5 4 1 3 4 4 4 4 8 (STOCK already made) Summary: [6 marks] Standard 1: Concentration _____________ How to prepare: Standard 2: Concentration _____________ How to prepare: Standard 3: Concentration _____________ How to prepare: Standard 4: Concentration _____________ How to prepare: Standard 5: Concentration _____________ How to prepare: 2. Suppose you collect the following data for the given BSA Standards. a) Fill in the absorbance column using the %T data. (Show one calc on the side) [2] 1/4 b) Create a standard curve graph, by hand (title, label axes (include units) and draw line of best fit) [5] c) Determine the concentration of the unknown from the given data and complete the table. [3] Sample % Transmittance BSA – 200 µg/mL 9.14 BSA – 100 µg/mL 23.4 BSA – 50 µg/mL 45.0 BSA – 25 µg/mL 66.8 BSA – 12.5 µg/mL 82.7 Blank 100 Unknown C 43.7 Unknown D 13.0 Absorbance Unknown Concentration ...
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