Unit 3: Laboratory Procedures

Today: 10/24/2013 Turn to page 45 in your lab manual and briefly read over the background section. Pay attention to Mass/Volume Concentration Equation For your background section include: A brief description of a spectrophotometer How this instrument works What it detects Define solvent and solute, and mention the most common solvent used in laboratories.

Today: 10/31/2013 Gummy Bear Sacrifice: Exothermic Reactions, Sugar Metabolism, Generally Awesome Flaming Jack: Boric Acid & Fluorescence Rock Candy: Super Saturation of a liquid using heat Dry Ice Bubbles: Surface tension & Sublimation

Today: 10/25/2013 Fill a small test tube with deionized water (do this now!). What is a spectrophotometer & how does it work? We will be testing our samples using a percent transmission setting on our specs. What does this mean? How many grams would it take to make 7 mL of solution at a concentration of 425 mg/ml? If I add three mL of water to the above solution, what is the new concentration of the solution?

Bellwork: 10/28/2013 What are the steps for using a spectrophotometer? What is the equation for determining mass/volume concentration when preparing a solution?

Today: 10/29/2013 You will complete the solution preparation & spec practice lab (pages 45-49). Clean up all the equipment that you used. Please use the sinks available in the Pre-AP Chem Lab For your data analysis section, compare your absorbance values with those of two other groups. Explain why they values are similar/different. For your conclusion: Answer the Thinking Like a Biotechnician questions on page 48. Answer the question posed in Procedure step #4. Include a brief description of possible errors. Answer the questions on page 49. Ignore 11, 13, 15, and 17.

Today: 10/30/2013 For the Background: Include all of the equations that you will need for this lab. Give an example of each equation being used in a practical setting. No “problem” is needed for this lab. The “safety” section needs to be detailed for this lab This lab will cover pages 50 – 54 in the lab notebook.

Today: 10/30/2013 Starting on page 51: Do the steps 1 and 2 for Part 1/2/3. If you are unable to confirm your math with another group, check with me. Make sure you show your work.

Bellwork: 11/01/2013 330 mL = _________ L _______________ 2. P-10/20/100/1000 measure what volumes?

Today: 11/01/2013 You must complete the following before the end of class, and your work must be initialed by me to receive full credit for the lab: Page 51 in the lab manual – Complete parts 1 and 2. Page 51 through 53 in the lab manual – Please read over part 3, 4, and 5 and draw the data tables for these parts in your lab notebook. Parts 3, 4, and 5 are not complicated procedures, but they will take most of the class on Monday to complete. I need a lot of help cleaning up my lab. If you finished today’s work, I would really appreciate the help cleaning up our lab space.

Today: 11/04/2013 How would you calculate the mass required to make the following solutions: 5 mL of 2.5% glucose solution: 2000 mg = ___________g 5L = ___________mL 6.3cm = _________mm

Today: 11/05/2013 Don’t wait for me to tell you to get started, get to work! You should be able to complete all of your lab work today. You will have time in class tomorrow to work on finishing your lab notebook work. **Addition for your lab instructions – The spec will be set at 540 nm, not 590 nm

Today: 11/04/2013 Changes to the lab: You will make two gelatin solutions of each concentration. You will run the experiment twice, once using your homemade Biuret Reagent, and a second time using a store-bought Biuret Reagent. For your conclusion you must answer all of the questions presented in the “Data Analysis/Conclusion” section of your lab manual, all “Thinking Like a Biotechnician” questions, and all math questions on page 54 except 11, 13, 15, 17, and 19.

Today: 11/07/2013 You need to complete the lab and the notebook portion of your lab today. I will collect your lab notebooks at the end of class! Changes to the lab: You will make two gelatin solutions of each concentration. You will run the experiment twice, once using your homemade Biuret Reagent, and a second time using a store-bought Biuret Reagent. For your conclusion you must answer all of the questions presented in the “Data Analysis/Conclusion” section of your lab manual, all “Thinking Like a Biotechnician” questions, and all math questions on page 54 except 11, 13, 15, 17, and 19. **Addition for your lab instructions – The spec will be set at 540 nm, not 590 nm

Using the Spec: Turn the instrument on and allow it to warm up for 15 minutes. Set your spec to the desired wavelength. Use a kim wipe to clean off any finger prints, oil, debris, etc., from the outside of your test tube. Insert the standard test tube (deionized water) and blank the spec. Insert your test sample and read/record both %transmittance and absorbance. Repeat steps 4 and 5 for each testable sample.

A) Metric System IS: International system of measurement Prefix Symbol Factor kilo k 1000 hecto h 100 (base unit) no prefix ---- 1 deci d 0.1 centi c 0.01 milli m 0.001 micro µ 0.000 001 (10-6) nano n 0.000 000 001 (10-9)

A) Metric System

A) Metric System To easily convert between prefixes we can use the BS rule

 *know these conversions!!!!! B) Measuring Volumes The main units we measure volumes in are microliters, milliliters and liters 1000 µL = 1 mL = 0.001 L  *know these conversions!!!!! Practice: 125 mL = _________ µL 33 mL = _________ L 4 L = _________ mL 200 µL = _________ mL 125,000 0.033 4,000 0.200

C) How do we measure volume in the lab? Graduated cylinder: Measure more than 10 mL Pipet: measure between 0.5 mL &10 mL Micropipet: less than 1 mL

C) How do we measure volume in the lab? Given the following volumes, what instrument would you use to measure that volume: 100 mL 25 mL 5 mL 10 mL 100 µL 25 µL 0.005 mL 56 mL Micropipet Graduated Cylinder Graduated Cylinder Micropipet Micropipet Pipet Pipet or Graduated Cylinder Graduated Cylinder

D) How to use a pipet Draw this: In order to use a pipet you much also have a pipet pump: Blue: 1- and 2-mL pipets Green: 5- and 10-mL pipets Red: 25- and 50-mL pipets

Roll the gear so the top raises, this will draw up the liquid into the pipet Roll the gear the opposite direction to expel the liquid DO NOT push the top of the pipet pump – this may break the pump When selecting a pipet to use remember to use the smallest pipet as possible to be as precise as possible

E) How to use a micropipet Used to measure amounts less than 1 mL 3 sizes of pipets: P10 0.5 µL to 10 µL P100 10 µL to 100 µL P1000 100 µL to 1000 µL Each micropipet has only a certain amount of precision Make sure you pay attention to this!

E) How to use a micropipet These are the readings for micropipets Main difference between 20 and 10 is the top limit, it still has the same unit places on the display

E) How to use a micropipet P10/P20

E) How to use a micropipet P100/P200

E) How to use a micropipet P1000/P2000

F) Making Solutions One of the most essential skills Labs often buy the solid form of a compound because it is often cheaper and many times multiple concentrations are used. Almost all reactions involving proteins and nucleic acids occur in an aqueous solutions. Solution contains 2 parts: Solute – what is being dissolved Solvent – what us doing the dissolving

F) Making Solutions To prepare a solution: Measure out solid mass of solute needed Add solid to container with a little water Dissolve solid Add enough water to reach desired volume

Today: 11/11/2013 We will review last week’s labs and discuss molarity Clean up day (hooray…)! Short metric conversion assignment

Common Units of Concentration G) Concentrations A measure of how many particles are in a certain volume Normality made be used for acids & bases only Common Units of Concentration Concentration Common Units of Measurement mass/volume g/L, mg/mL, µg/mL, µg/µL % mass % Molarity M (mol/L), mM (mmol/L), µM (µmol/L)

H) Preparing Mass/Volume Concentration Mass/Volume Concentration Equation Conc. desired x volume desired = mass of solute (g/mL) (mL) (g) Example: How do you prepare 100 mL of 0.05 g/mL solution of hemoglobin? Use the equation to determine mass of solute. 0.05 g/mL x 100 mL = 5 g hemoglobin Give directions for making solution: Measure out 5 g of hemoglobin and pace into a 100 mL flask, add about 10-20 mL of dH2O. Dissolve all hemoglobin then add enough water to make 100 mL of solution.

H) Preparing Mass/Volume Concentration Mass/Volume Concentration Equation Conc. desired x volume desired = mass of solute (g/mL) (mL) (g) Practice: How do you prepare 150 mL of 2 mg/mL gelatin? Use the equation to determine mass of solute. 2 mg/mL x 150 mL = 300 mg gelatin = 0.3 g gelatin Give directions for making solution: Measure out 0.3 g of gelatin and dissolve in a little bit of water. Add enough water to make 150 mL of solution.

I) Preparing % Mass/Volume Concentration % Mass/Volume Concentration Equation Convert % to decimal Decimal % x volume desired = mass of solute (g/mL) (mL) (g) Example: How do you prepare 50 mL of 10% NaCl solution? Use the equation to determine mass of solute. 10% → 0.10 g/mL 0.10 g/mL x 50 mL = 5 g NaCl Give directions for making solution: Measure out 5 g of NaCl and dissolve in a small amount of water. Add enough water to make 50 mL of solution.

I) Preparing % Mass/Volume Concentration % Mass/Volume Concentration Equation Convert % to decimal Decimal % x volume desired = mass of solute (g/mL) (mL) (g) Example: How do you prepare 60 mL of 10% glucose? Use the equation to determine mass of solute. 10% → 0.10 g/mL 0.10 g/mL x 60 mL = 6 g glucose Give directions for making solution: Measure out 6 g of glucose and dissolve in a small amount of water. Add enough water to make 60 mL of solution.

J) Preparing solutions using Molarity Molarity Units: mol/L mole = the number of particles in a substance that is equal to the mass (in grams) on the periodic table Ex: 1 mole NaCl = _____ g 1 mol of any substance will not be the same as a different substance! 58.5 1 mole Na = 23.0 g 1 mole Cl = 35.5 g 1 mole NaCl = 23.0 g + 35.5 g = 58.5 g

J) Preparing solutions using Molarity Practice: Solve the following. 1 mol C6H12O6 = ________ g 1 mol CaCl2 = ________ g 180.0g and 111.1g are called the molecular weight (MW) or formula weight of its substance 180.0 (12.0 x 6) + (1.0 x 12) + (16.0 x 6) 111.1 (40.1 x 1) + (35.5 x 2)

J) Preparing solutions using Molarity Molarity Concentration Equation Volume wanted x molarity x MW = mass of solute (L) (mol/L) (g/mol) (g) Example: How many grams are needed to make a 75 mL solution of 2M CaCl2? 0.075 L x 2M x 111.1 g/mol = 16.67g CaCl2 Practice: How do you prepare 20 mL solution of 0.25 M NaCl? 0.020 L x 0.25M x 58.5 g/mol = 0.293 g NaCl Measure out 0.293g of NaCl and dissolve in enough water to make 20 mL of solution.

Bellwork: 11/12/2013 What mass of calcium chloride (CaCl2) is needed to make 125 mL of 0.55 M CaCl2 solution?

Bellwork: 11/12/2013 What mass of calcium chloride (CaCl2) is needed to make 125 mL of 0.55 M CaCl2 solution? 1st convert 125 mL into L (molarity = moles/liter) 125 mL = 0.125 L Then use the molarity concentration equation

Volume wanted x molarity x MW = mass of solute Bellwork: 11/12/2013 Volume wanted x molarity x MW = mass of solute 0.125 L x 0.55 M CaCl2 x 111 grams/mole = 7.63 grams of CaCl2 and add solvent up to 125 mL

K) Dilutions Buying concentrated solutions can save money If you need a very dilute solution starting from a more concentrated solutions may be required

K) Dilutions Dilution Equation C1V1=C2V2 C = Concentration V = Volume 1 & 2 stand for different solutions Concentrations can be in any unit as long as both are the same, same with volume Example: How would you prepare 1 L of 1mg/mL protein solution from 100mg/mL concentrated stock? C1= 1 mg/mL V1=1L C2=100 mg/mL V2=? (1 mg/mL)(1L) = (100 mg/mL)(V2) V2=10mL Measure out 10 mL of protein stock solution and mix in enough water to make 1 L of solution.

K) Dilutions Dilution Equation C1V1=C2V2 C = Concentration V = Volume 1 & 2 stand for different solutions Concentrations can be in any unit as long as both are the same, same with volume Practice: How do you make 200 mL of 10mM CaCl2 from 75mM CaCl2 solution? C1= 10 mM V1= 200 mL C2=75 mM V2=? (10 mM)(200mL) = (75 mM)(V2) V2=26.6mL Measure out 26.6 mL of CaCl2 and mix in enough water to make 200 mL of solution.

Get to Work! Changes to the Lab: Step 3 (page 56) – You do NOT need to graph your data. The trend(s) should be obvious through a data chart. For your conclusion: Possible errors Results with evidence Practical use for this technique Thinking like a Biotechnician, questions 1, 2, 3 A summary of page 58 – why is pH biologically important? Etc.

Get to Work! For your background section: Explain: mole, molarity, Avogadro's number, and molecular weight Write out the molarity concentration equation (include all units) Solve the following, showing all of your work: What is the molarity of a solution containing 9.478 grams of RuCl3 in enough water to make 1.00 L of solution? How many grams of KMnO4 are required to prepare 1.0 L of a solution of 1.5 M KMnO4?

Get to Work! For your background section: Explain: mole, molarity, Avogadro's number, and molecular weight Write out the molarity concentration equation (include all units) Solve the following, showing all of your work: What is the molarity of a solution containing 9.478 grams of RuCl3 in enough water to make 1.00 L of solution? How many grams of KMnO4 are required to prepare 1.0 L of a solution of 1.5 M KMnO4?

Today: 11/15/2013 You must: Complete Lab 3g, pages 55-57 Clean Up your equipment from yesterday (everything can go down our lab sink) Turn in your lab notebook before you leave

Today: 11/18/2013 Lab 3h (page 59) Plan on taking the exam for this unit on Friday. The exam will focus on pages 31 – 62 in the lab manual. The lab will consist of written and practical questions.

Today: 11/19/2013 Brief information about AAI National Conference (immunology2014.org) Some groups still need to clean up from last week A change in pace of labs You should be able to have about half of your solutions made today. Don’t hog the analytical balances.

Today: 11/18/2013 Lab 3h (page 59) Background expectations: What does it mean to dilute a solution. Why would this be necessary in a laboratory setting? Write out & define the dilution equation What is a serial dilution? What type experiment would use a serial dilution? Showing all your work, answer the following questions: Prepare 200 mL of 25 mg/mL NaOH solution from a 150 mg/mL NaOH 800 mL of 5X CuSO4 .5 H2O solution from 25X CuSO4 .5 H2O

Today: 11/21/2013 Address the cloudy solution issue Everyone should have at least one set of absorbance data to graph Only one graph will be required for your lab notebook Hand out exam review sheets Unit #3 Exam will be one Monday Lab notebooks will be turned in Friday for grading (hopefully) Unit math/practical quiz will be on Friday Turn in your AAI information sheets if you are interested in going on the trip in May

Today: 11/21/2013 Changes to the lab & additions for conclusions: Only one graph will be required (make sure all axis are labeled) Do NOT do any of the “Diagram of How to Prepare It” questions (2, 4, 6, 8, 10, 12, 14) Complete all Thinking Like a Biotechnician Questions Explain your results, including the cloudiness issue, as well as what you would have predicted to see in the absorbance without the cloudiness problem Possible error section No Practical application section is needed, this will be answered through the Thinking Like a Biotechnician Questions Add: Do you think it would be beneficial to have this unit as our first unit, including labs I skipped from pages 31-40. Explain why or why not

Unit Equations: % Mass/Volume Concentration Equation Conc. desired x volume desired = mass of solute (g/mL) (mL) (g) % Mass/Volume Concentration Equation Convert % to decimal Decimal % x volume desired = mass of solute

Unit Equations: Molarity Concentration Equation Volume wanted x molarity x MW = mass of solute (L) (mol/L) (g/mol) (g) Dilution Equation C1V1=C2V2 C = Concentration V = Volume 1 & 2 stand for different solutions Concentrations can be in any unit as long as both are the same, same with volume