Medical Biochemistry I Lab Exercise 1

Course Information

• Course name: Medical Biochemistry I
• Course code: MD100
• Lab Exercise 1 title: Introduction to Techniques - Use of Pipetman, Sample Preparation and Dilutions
• Semester: Fall 2024
• Instructors: Andreou M & Kyriakou TC
• University: European University Cyprus
• School: School of Medicine

Objectives

• Introduction: Lab characteristics/guidelines
• Part A: Use of Pipetman (theoretical background and exercises)
• Part B: Sample Preparation (main units of concentration and exercises)
• Part C: Dilutions (dilution formula and exercises)

COVID-19 Safety Protocol

• Be on time to avoid crowds at the lab entrance.
• Carry all protective gear (lab coat with cuffed sleeves, mask, goggles).
• Students without protective gear are not allowed in the lab.
• Practice hand hygiene (wash hands with soap, then gloves).
• Disinfect working surfaces before and after experiments.
• Report any symptoms (fever, cough, breathing difficulties) immediately.

Absences

• Mandatory participation: physical presence or synchronous participation required (permission from the department).
• Excused absences: only two justified absences allowed (severe health condition or emergency).
• Announcements: justified absences must be announced before the beginning of the class.
• Repeating experiments: Lab experiments are not repeatable.

Preparation

• Study uploaded materials.
• Print the protocol/experimental methodology.
• Mark materials/equipment needed on the protocol.
• Mental preparation for demanding applications.
• Required attire: Closed-toe shoes, lab coat, goggles, gloves, hair tied back (ponytail).
• No food or drinks in the lab.

Lab Notebook

• Record all information and explanations.
• Note down protocol applications with dates.
• Calculate reagent concentrations (e.g., C1V1=C2V2).
• Evaluate results and identify areas for improvement.

Lab Etiquette

• Move cautiously (like in a bubble).
• Be aware of your surroundings.
• Work collaboratively as a team.
• Raise your hand to ask questions or for clarification.
• Avoid sudden or rushed movements.
• Work quietly and respectfully.

Contact Information

• Email addresses: [email protected] & [email protected]
• Office location: N30 (2nd floor - Medical Building)
• Email policy: Students must use their EUC email when contacting faculty members.

Materials and Equipment

• Pipetman (P1000, P200, P10)
• Pipet tips
• Distilled Water
• 1.0 M CuSO4 solution
• Cuvettes
• Balance machine
• Spectrophotometer
• Weighing boats
• Waste containers

Pipetman Use

• For volumes less than 1 mL, Pipetman is the most common method.
• Pipettes are basic tools in a fully equipped laboratory.
• Types of Pipetman:
  • P1000 (200-1000 μL)
  • P100 (10-100 μL)
  • P10 (0.5-10 μL)
• 1L = 1000 mL and 1 mL = 1000 μL

Pipetman Procedure (Diagram-based)

• Illustration of different steps in using a Pipetman (A-E).
• Diagram displays steps: upper stop, first stop, second stop.
• Illustrations show important measurements (1 cm, 30 degrees).

Part A - Pipetman Accuracy

• Accuracy checking method: weighing water.
• Water density at room temperature: 1 g/mL
• Each microliter (μL) should weigh 0.001 g (water).
• Accurate pipette volume delivery equals water weight.

Part A – 1000 μL Pipetman Exercise

• Instructions to accurately weigh different volumes of distilled water (1000 μL, 650 μL, 358 μL).
• Table to record readings and calculate averages.

Part A – 200 μL Pipetman Exercise

• Instructions for accurately weighing various volumes of distilled water (200 μL, 65 μL, 47.5 μL).
• Table for recording readings and calculating averages.

Part B – Sample Preparation

• Solvent: the liquid that makes up most of a solution.
• Aqueous solution: a solution using water as a solvent.
• Solute: the substance dissolved in the solvent.
• Definitions: Solution concentration = amount of solute / amount of solution.

Concentration Units

• % by mass (mass/mass)
• % by volume (volume/volume)
• % weight/volume (mass/volume)
• Molarity (M)

Part B – Example 1: % by Mass

• Calculation example to determine the % by mass concentration of a 100 g solution, where 5 g of NaOH is dissolved in water.
• Formula for calculation presented (g solute/g solution * 100%).

Part C – Dilutions

• Many biochemical solutions are prepared by diluting a concentrated stock solution.
• Dilutions require consideration of both the final desired concentration and volume of the diluted material.
• Important formula: C1V1 = C2V2
• Definitions:
  • C1: Initial/stock solution concentration
  • V1: Volume of the stock solution
  • C2: Desired final solution concentration
  • V2: Desired final solution volume
• Parallel dilutions: Using the same equation to dilute solutions with known concentration factors (eg 10X, 20X, 100X).

Part C – Parallel Dilution Example

• Example calculating the volume needed to prepare 50 mL of 1X NaOH from a 10X NaOH stock solution.
• Dilution factors indicated by numerical suffixes like 0.5X, 1X, 2X.