2. Ellyn Daugherty SM Biotech Career Pathway www.BiotechEd.com www.emcp.com/biotech www.sargentwelch.com/biotech aeedaugher@aol.com Biotech 1 SLOP Text - Chapter 3 Lab Manual - Chapter 3 Student CD Tutorials Course Planner Overcoming Molephobicity (5 Steps to Teach Solution Prep)

3. 1.What is a solution? 2.Solvent Measurement 3.Measure the Solute 4.Mix the Solute and Solvent 5.Check the Solution Preparation Five Steps to Teach Solution Prep

4. Solutes (usually measured in g) Solvents (usually measured in mL) Mixed together until solute dissolves in solvent Described by the portion of solute to solution volume usually in one of 3 ways: - mass/volume - % mass/volume - molarity Step 1. What is a Solution ? 8 mg/mL4 mg/mL2 mg/mL concentration = solute solution volume 2% glucose solution 10% sodium hydroxide solution 5 g/L protease 175 mg/mL rennin 25 µg/µL chymosin 1M NaOH 50 mM TRIS 5 µM CaCl 2

5. Volume of solvent is usually measured using one of 3 instruments. For each, ask: What is the total volume that can be measured? What is the value of each type of graduation? Step #2 Solvent Measurement graduated cylinder pipet micropipet

6. L mL µL Volume and Mass Measurement requires Metric Measurements and Conversions 1000 92 mL = _________ µL357 µL = _________ mL 0.5 g = _________ mg0.557 mg = _________ g

7. Step #3 Solute Measurement Solutes are usually dry chemicals and their mass is measured on a scale/balance and reported in grams (g). For a specified concentration of a specified volume of solution, the technician must determine the amount of mass of solute needed for the volume of solution at some final concentration. 3 different equations, one for each concentration units group - mass/volume - % mass/volume - molarity

8. Determine the Mass of Solute (g) 3 different equations, one for each concentration units group mass/volume % mass/volume molarity ___________g/mL ____________mL _________________ concentration desired X volume desired = mass of solute needed (g) __________g/mL ____________mL _________________ conc desired (decimal) X volume desired = mass of solute needed (g) _________ ____________ __________ __________ concentration X volume desired X molecular weight = mass of solute needed desired (mol/L) (L) (g/mol) (g)

9. Measuring for Mass/Volume Solutions To make 100 mL of a 5 mg/mL hemoglobin solution ___________ _____________ ___________g______ concentration desired X volume desired = mass of solute to be weighed 5 mg/mL100 mL500 mg = 0.5 g

10. Measuring for % Mass/Volume Solutions __________ ____________ ____________g_____ conc desired (decimal) X volume desired = mass of solute needed To make 100 mL of a 3% CuSO 4 solution 3 % = 0.03 conc desired X volume desired = mass of solute needed 3 g CuSO 4 0.03 g/mL100 mL

11. Measuring for Molar Solutions To make 300 mL of a 0.5M NaOH solution. Convert 300 mL = 0.3 L To make 300 mL of a 0.5M NaOH solution. Convert 300 mL = 0.3 L _________ ____________ __________ _________g__ concentration X volume desired X molecular weight = mass of solute desired (mol/L) (L) (g/mol) (g) Molecular Weight ? Moles are used to count molecules and atoms since they are too small to count individually. 1 mole equals the amount of molecules that gives a mass, in grams, equal to that substance’s molecular or formula weight. The formula weight (FW or MW) can be determined using a periodic table or by looking at a chemical reagent bottle. Molecular Weight ? Moles are used to count molecules and atoms since they are too small to count individually. 1 mole equals the amount of molecules that gives a mass, in grams, equal to that substance’s molecular or formula weight. The formula weight (FW or MW) can be determined using a periodic table or by looking at a chemical reagent bottle.

12. For Molar Solutions you need the MW Determine the molecular weight of the solute The formula weight can be determined using a periodic table or by looking at a chemical reagent bottle Determine the molecular weight of the solute The formula weight can be determined using a periodic table or by looking at a chemical reagent bottle

13. Measuring for Molar Solutions To make 300 mL of a 0.5M NaOH solution. Convert 300 mL = 0.3 L To make 300 mL of a 0.5M NaOH solution. Convert 300 mL = 0.3 L 6 g of NaOH is measured on a balance and put into an appropriate vessel. Then, solvent is added up to the 300 mL graduation. concentration X volume desired X molecular weight = mass of solute desired (mol/L) (L) (g/mol) (g) 0.5 mol/L0.3 L 40 g/mol6 g

14. 4. Mixing the Solute and Solvent Mass/Volume solutions 100 mL of a 5 mg/mL hemoglobin solution. solvent (dH 2 0 or buffer) 0.5 g hemoglobin 100 mL 5 mg/mL 100 mL 500 mg = 0.5 g concentration desired X volume desired = mass of solute to be weighed 250 mL beaker

15. PPT Demos on the Encore® CD 25 tutorials for the basic SLOP (standard lab operating procedures) skills

16. Step 5 Checking the Solution Preparation I like to teach solution prep using copper sulfate since it is blue and can be visualized (relative number of molecules compared) in the Spec 20D+ at 600 nm) I also check the volume and labels on all tubes. Other methods of checking solution prep?

17. Now YOU try it. Make the following solutions but before you do: Do all the calculations showing the equations with all the units of measurement shown. Diagram how you are going to prepare the solution. After you prepare the solution, check it in the spec. Demonstrate how to take up and dispense a sample. Make all the solutions in 15 mL tubes and then transfer 3 mL into 13x100 mm tubes for checking in the spec. 4 mL of 0.5 M cupric sulfate pentahydrate 5 mL of 250 mM cupric sulfate pentahydrate 6 mL 125 mM cupric sulfate pentahydrate Check each solution in the spec at 600 nm by measuring the absorbance. Use 3 mL of the solution to be checked, in a 13x100 mm cuvette. Note: M mM 1000