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Questions and Answers
What is the primary active reagent used in the glucose oxidase method?
What is the primary active reagent used in the glucose oxidase method?
What color change indicates a presence of glucose in urine during a dipstick test?
What color change indicates a presence of glucose in urine during a dipstick test?
What does the term 'renal threshold for glucose' refer to?
What does the term 'renal threshold for glucose' refer to?
What is typically analyzed with a glucometer to measure blood glucose levels?
What is typically analyzed with a glucometer to measure blood glucose levels?
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Which of the following is a possible outcome if blood glucose levels exceed 140 mg/dl?
Which of the following is a possible outcome if blood glucose levels exceed 140 mg/dl?
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Which laboratory technique involves immersion of test strips in urine samples?
Which laboratory technique involves immersion of test strips in urine samples?
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What is the expected result of a normal urine test concerning glucose presence?
What is the expected result of a normal urine test concerning glucose presence?
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During the colorimetric analysis of glucose, what do the colored products formed indicate?
During the colorimetric analysis of glucose, what do the colored products formed indicate?
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What is the primary purpose of specimens taken from the body?
What is the primary purpose of specimens taken from the body?
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Which of the following statements accurately describes plasma?
Which of the following statements accurately describes plasma?
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What is the first step in plasma separation from a whole blood sample?
What is the first step in plasma separation from a whole blood sample?
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What occurs during the serum separation process?
What occurs during the serum separation process?
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Which specimen would most likely be used to test respiratory conditions?
Which specimen would most likely be used to test respiratory conditions?
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What is included in whole blood specimens?
What is included in whole blood specimens?
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Which body fluid is NOT typically classified as a specimen for laboratory testing?
Which body fluid is NOT typically classified as a specimen for laboratory testing?
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What is the difference between serum and plasma?
What is the difference between serum and plasma?
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What is the primary purpose of using a capillary sample?
What is the primary purpose of using a capillary sample?
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Which vein is not commonly selected for venipuncture at the cubital fossa?
Which vein is not commonly selected for venipuncture at the cubital fossa?
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What is the correct order of steps for the blood sampling procedure?
What is the correct order of steps for the blood sampling procedure?
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Which of the following tests does NOT require fasting?
Which of the following tests does NOT require fasting?
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What is a recommended precaution regarding patient positioning during a blood test?
What is a recommended precaution regarding patient positioning during a blood test?
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Which among the following is NOT a part of the required equipment for sampling?
Which among the following is NOT a part of the required equipment for sampling?
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For how long should a patient typically fast before tests like fasting blood glucose?
For how long should a patient typically fast before tests like fasting blood glucose?
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What role does the tourniquet serve during venipuncture?
What role does the tourniquet serve during venipuncture?
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What is the formula for calculating molarity?
What is the formula for calculating molarity?
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How is molality defined?
How is molality defined?
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What does equivalent weight depend on?
What does equivalent weight depend on?
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If 2 moles of NaCl are dissolved in 1 liter of water, what is the molarity of the resulting solution?
If 2 moles of NaCl are dissolved in 1 liter of water, what is the molarity of the resulting solution?
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What is the result when you dissolve 3 moles of KOH in 3 liters of water?
What is the result when you dissolve 3 moles of KOH in 3 liters of water?
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Which statement is true regarding normal solution?
Which statement is true regarding normal solution?
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Which of the following methods is correct for preparing a solution of a specified concentration?
Which of the following methods is correct for preparing a solution of a specified concentration?
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What is meant by the term 'valence' in chemistry?
What is meant by the term 'valence' in chemistry?
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What is the primary enzyme used in glucometers to measure glucose?
What is the primary enzyme used in glucometers to measure glucose?
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Which of the following is NOT a requirement for using a glucometer?
Which of the following is NOT a requirement for using a glucometer?
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What is the purpose of the alcohol swab in the glucometer testing process?
What is the purpose of the alcohol swab in the glucometer testing process?
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In the colorimetric method for measuring glucose, what is produced as a byproduct of glucose oxidation?
In the colorimetric method for measuring glucose, what is produced as a byproduct of glucose oxidation?
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How does the measurement of glucose concentration typically occur in a colorimetric method?
How does the measurement of glucose concentration typically occur in a colorimetric method?
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What is the role of o-toluidine in the glucose oxidase colorimetric method?
What is the role of o-toluidine in the glucose oxidase colorimetric method?
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What should be done after obtaining the blood sample in the glucometer testing process?
What should be done after obtaining the blood sample in the glucometer testing process?
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What are the colored products in the glucose oxidase method indicative of?
What are the colored products in the glucose oxidase method indicative of?
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What is the normal range for total plasma protein concentration?
What is the normal range for total plasma protein concentration?
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Which statement about the Biuret test is true?
Which statement about the Biuret test is true?
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What is the main principle behind protein analysis in body fluids?
What is the main principle behind protein analysis in body fluids?
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Which plasma protein constitutes the largest percentage in normal plasma?
Which plasma protein constitutes the largest percentage in normal plasma?
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In an alkaline medium, how do proteins migrate during electrophoresis?
In an alkaline medium, how do proteins migrate during electrophoresis?
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How many fractions of plasma proteins can be distinguished by their mobility?
How many fractions of plasma proteins can be distinguished by their mobility?
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What is a limitation of protein detection in urine or cerebrospinal fluid?
What is a limitation of protein detection in urine or cerebrospinal fluid?
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What does the intensity of the violet color in the Cu2+ protein complex indicate?
What does the intensity of the violet color in the Cu2+ protein complex indicate?
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Which of the following is NOT a type of hazard found in the laboratory?
Which of the following is NOT a type of hazard found in the laboratory?
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What safety measure must be followed when handling broken glassware?
What safety measure must be followed when handling broken glassware?
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Which action is appropriate when a cut occurs during laboratory work?
Which action is appropriate when a cut occurs during laboratory work?
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Which statement best describes the proper use of heating equipment in the laboratory?
Which statement best describes the proper use of heating equipment in the laboratory?
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What is the first step in responding to a burn in the laboratory?
What is the first step in responding to a burn in the laboratory?
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When should you wash your hands during laboratory procedures?
When should you wash your hands during laboratory procedures?
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Which condition must be met before heating glassware?
Which condition must be met before heating glassware?
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What should you do if a chemical spill occurs during an experiment?
What should you do if a chemical spill occurs during an experiment?
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What is the significance of the renal threshold for glucose in human physiology?
What is the significance of the renal threshold for glucose in human physiology?
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During the glucose oxidase method, what role does hydrogen peroxide play?
During the glucose oxidase method, what role does hydrogen peroxide play?
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Which component is NOT part of the glucometer testing process?
Which component is NOT part of the glucometer testing process?
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What is the purpose of comparing the test strip color against a color chart after a urine test?
What is the purpose of comparing the test strip color against a color chart after a urine test?
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Which of the following statements accurately describes the function of Peroxidase in the glucose oxidase method?
Which of the following statements accurately describes the function of Peroxidase in the glucose oxidase method?
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To perform a urine test using a dipstick method accurately, what is a critical initial step?
To perform a urine test using a dipstick method accurately, what is a critical initial step?
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What is the typical glucose concentration threshold at which glucose begins to appear in urine?
What is the typical glucose concentration threshold at which glucose begins to appear in urine?
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What factor is essential for the accuracy of readings obtained from a glucometer?
What factor is essential for the accuracy of readings obtained from a glucometer?
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What would be the correct action to prepare 1 N NaOH solution?
What would be the correct action to prepare 1 N NaOH solution?
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How is osmotic pressure related to molar concentration in a solution?
How is osmotic pressure related to molar concentration in a solution?
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What is the equivalent weight of NaOH?
What is the equivalent weight of NaOH?
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What does osmolarity measure in a solution?
What does osmolarity measure in a solution?
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What will happen to RBCs in an isotonic solution?
What will happen to RBCs in an isotonic solution?
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Which of these describes the significance of using isotonic solutions in medical applications?
Which of these describes the significance of using isotonic solutions in medical applications?
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What is the molar mass of NaCl?
What is the molar mass of NaCl?
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What is the primary implication of osmosis in medical scenarios?
What is the primary implication of osmosis in medical scenarios?
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Which specimen is least likely to be used for biochemical analysis in a laboratory setting?
Which specimen is least likely to be used for biochemical analysis in a laboratory setting?
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What is the primary difference in the collection process between obtaining plasma and serum?
What is the primary difference in the collection process between obtaining plasma and serum?
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In the context of blood specimens, what are the cellular components of whole blood primarily composed of?
In the context of blood specimens, what are the cellular components of whole blood primarily composed of?
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Why is it essential not to rub the eye if a foreign object is lodged in it?
Why is it essential not to rub the eye if a foreign object is lodged in it?
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Which body fluid type is not typically involved in biochemical laboratory tests?
Which body fluid type is not typically involved in biochemical laboratory tests?
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What is the primary action performed to separate serum from whole blood?
What is the primary action performed to separate serum from whole blood?
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Which of the following specimens must contain anticoagulant during collection to obtain plasma?
Which of the following specimens must contain anticoagulant during collection to obtain plasma?
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What is the most significant purpose of collecting specimens like blood and urine in laboratory settings?
What is the most significant purpose of collecting specimens like blood and urine in laboratory settings?
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Which type of molecule has an uneven distribution of electrical charge resulting in a dipole?
Which type of molecule has an uneven distribution of electrical charge resulting in a dipole?
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What defines a concentrated solution?
What defines a concentrated solution?
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What size range of solute particles is classified as visible to the naked eye?
What size range of solute particles is classified as visible to the naked eye?
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Which of the following substances is a universal solvent?
Which of the following substances is a universal solvent?
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What is the correct definition of molarity?
What is the correct definition of molarity?
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Which substance is an example of a nonpolar solvent that dissolves nonpolar solutes?
Which substance is an example of a nonpolar solvent that dissolves nonpolar solutes?
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Which term describes a solution that has a low concentration of solute dissolved?
Which term describes a solution that has a low concentration of solute dissolved?
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What measurement expresses the number of moles of solute present in one kilogram of solution?
What measurement expresses the number of moles of solute present in one kilogram of solution?
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What is the significance of the copper-protein complex formed in protein analysis?
What is the significance of the copper-protein complex formed in protein analysis?
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Which plasma protein accounts for 50% to 60% of total plasma protein concentration?
Which plasma protein accounts for 50% to 60% of total plasma protein concentration?
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During the Biuret test, at which wavelength is the absorption measured?
During the Biuret test, at which wavelength is the absorption measured?
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What is a primary limitation when detecting proteins in urine or cerebrospinal fluid?
What is a primary limitation when detecting proteins in urine or cerebrospinal fluid?
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What does protein mobility during electrophoresis depend on?
What does protein mobility during electrophoresis depend on?
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Which fraction of plasma proteins is least likely to migrate towards the anode during electrophoresis?
Which fraction of plasma proteins is least likely to migrate towards the anode during electrophoresis?
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How is the measurement of total plasma protein typically expressed?
How is the measurement of total plasma protein typically expressed?
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What characteristic of the Cu2+ binding process is essential for protein quantification?
What characteristic of the Cu2+ binding process is essential for protein quantification?
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Study Notes
Biochemistry Laboratory Safety
- Hazards: Exposure to blood, body fluids, specimens (bacteria, viruses), acids, alkalis, toxic chemicals, radioactive waste disposal, fire, and electrical hazards.
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General Safety Rules:
- Carefully read all instructions before starting any experiment.
- Wear appropriate personal protective equipment (PPE): safety goggles, lab coat, gloves, face mask, and hair cap.
- Wash hands thoroughly with soap and water after handling chemicals.
- Avoid contact with chemicals by keeping hands away from the face.
- Do not mix chemicals unless instructed.
- Notify the teacher about any spills or accidents.
- Clean up the lab area after the experiment.
Glassware Safety
- Broken Glassware Disposal: Dispose of broken glassware in a special glass disposal container.
- Pouring Liquids: When pouring liquids into glassware, keep the container at least a hand's breadth from the edge of the table.
- Broken Glassware: Notify the teacher immediately if glassware breaks.
Heating Safety
- Handling Hot Objects: Use protective gloves when handling hot objects.
- Open Flames: Never reach across an open flame.
- Heating Test Tubes: When heating test tubes, move them slowly over the flame to distribute heat evenly and avoid pointing them at people.
- Heating Glassware: Only heat thoroughly dry glassware. Heat glassware by placing it on a wire gauze platform on a ring stand. Do not hold it in your hand.
First Aid
- Burns: Immediately flush with cool water.
- Cuts & Bruises: Do not touch an open wound without safety gloves; press directly on minor cuts to stop bleeding; apply cold compress to reduce swelling.
- Eyes: Flush eyes immediately with plenty of water for several minutes. Do not rub the eye if a foreign object is lodged in it.
Sampling
- Specimens: A specimen is any substance taken from the body for testing. Common specimens include blood, urine, stool, CSF, saliva, semen, respiratory secretions, and other body fluids.
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Blood Specimens:
- Whole blood: Includes all cellular components (RBCs, WBCs, platelets) and fluid components.
- Plasma: The liquid portion of blood without clotting factors.
- Serum: The liquid portion of blood without clotting factors.
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Plasma and Serum Separation:
- Plasma: Anticoagulants are used to prevent blood clotting. The blood sample is then centrifuged to separate the cells (blood cells) from the liquid portion (plasma).
- Serum: Samples are left undisturbed at room temperature for 15-30 minutes to allow clotting. Then the sample is centrifuged to separate the solid clot from the liquid serum.
- When to Use Whole Blood, Plasma, or Serum:
- Whole Blood: Used for blood grouping, blood cultures, and hemoglobin tests
- Plasma: Used for coagulation tests and clotting factor analysis.
- Serum: Used for creatinine, glucose, and lipid profiles.
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Types of Collecting Tubes:
- Plain tubes: No anticoagulant.
- Serum separator tubes (SST): Contains clot activator and gel separator.
- Anticoagulant-containing tubes: Citrate, EDTA, heparin, sodium citrate, and sodium fluoride. The choice depends on the analysis.
- Labeling Tubes: Tubes must be labeled with patient information (first and last name, ID number, date and time of collection, collector's initials, and additional information like fasting).
Lab Tests Requiring Fasting
- Fasting blood glucose, HOMA-IR, C-peptide, lipid profile (triglycerides), iron tests, Vit B12, Gamma glutamyl transferase.
- Fasting means no food or drinks (except water) for 8-12 hours.
Procedure Steps for Blood Collection
- Sensitization: Hand washing and applying gloves.
- Tourniquet Application Place the tourniquet 3 to 4 inches above the venipuncture site.
- Site Cleanse: Clean the site with a gauze pad soaked in 70% isopropyl alcohol or a commercially prepared alcohol prep pad.
- Equipment Preparation: Select a syringe and needle of appropriate size.
- Tourniquet Reapplication and Needle Insertion: Reapply the tourniquet and insert the needle at a 15-30° angle.
- Needle Removal: Withdraw the needle while applying pressure. Apply an adhesive bandage or tape
- Blood Transfer to Collecting Tube: Transfer prepared blood into the tube according to requirements.
- Collecting Tube Labeling: Label the collection tube immediately, including patient information and appropriate information. -Sample Transport and Processing: Transport the samples immediately to the lab, using appropriate cold storage if needed, to preserve specimen integrity.
Complications of Blood Collection
- Fainting, Hematoma, Petechiae, Bleeding, Seizures.
Solutions and Osmolarity
- Solutions: Homogenous (true solutions: crystallloid), Heterogeneous (suspensions, colloids).
- Colloids: Particles with sizes ranging from 1 to 1000 nm, visible through a microscope.
- Suspensions: Particles with sizes greater than 1000 nm, visible to the naked eye.
- Solute Material dissolved to form a solution.
- Solvent The liquid dissolving medium in a solution.
- Osmolarity: Number of solute particles per liter of solvent.
- Osmolarity Formula: Moles solute / Liter solution
- Osmosis: Diffusion of a solvent across a semipermeable membrane from an area of lower solute concentration to an area of higher solute concentration.
- Isotonic: A solution having the same osmotic pressure as the fluid it is placed in (no net movement of water in or out of the cell).
- Hypotonic: A lower osmotic pressure than the surrounding fluid, water moves into the cell.
- Hypertonic: A higher osmotic pressure than the surrounding fluid, water moves out of the cell
- Clinical Relevance of Osmotic Effects: Solutions are often made isotonic to prevent irritation and damage to delicate membranes and/or tissues.
- Cholera: Overpopulation of the intestines reverses the flow of water absorption, resulting in diarrhea and dehydration, due to the agent Vibrio cholerae.
Glucose Measurement Methods
- Glucose is a simple sugar. The terms glucose and sugar are frequently used interchangeably when discussing glucose levels in body fluids.
- Glucose Oxidase Method: A highly specific method for measuring glucose levels. It involves a double enzymatic reaction in the presence of an indicator.
- Glucometers: A device that analyzes capillary blood samples to provide numerical readings of glucose levels in the blood.
Urine Test Strips
- Components: Urine test strips use a thin strip of plastic or paper impregnated with chemicals to detect the presence of different compounds. This method produces colored results that can be compared to a chart to determine glucose levels, and other substances.
- Steps: Immerse the test strip in a well-mixed urine sample for a short period (1–2 minutes), remove excess urine and leave the strip undisturbed until the desired color changes are produced, and compare the color changes to a printed color chart to determine the concentration of the substance.
Glucometer
- Purpose: To determine the amount of glucose in blood, typically from a finger prick.
- Process: Device analyzes capillary blood sample and provides a numerical reading for glucose concentration.
Principle of Glucometers
- Use test strips with glucose oxidase that react with the glucose in the blood sample and a sensor to determine the glucose level.
Colorimeter
- Purpose: Used in clinical labs to measure the concentration of colored compounds by measuring their absorbance, at a specific wavelength of light.
- Colorometer Process: The method involves adding reagents to a sample that produces colored products. The intensity of the color is related to the concentration of the substance being measured. This measurement is then read with the device.
Protein Measurement in Body Fluids
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Biuret Method: Determines protein concentration based on the binding of copper ions to peptide bonds in an alkaline solution, producing a violet colored complex. The intensity of the color is proportional to protein concentration. This method is effective for measuring protein in a wide range of fluids, except in low concentrations.
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Plasma Protein Measurement: Used to measure total protein and specific types like albumin, globulins, and fibrinogen.
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Normal Ranges: Plasma protein concentration should be between 6.0 and 8.0 g/dL. Normal plasma albumin ranges from 3.5 to 5.5 g/dL and other globulins are also measured at the same time.
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Colorimetric Assays: A traditional method to measure protein concentration in fluids using colorimetric reactions.
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Electrophoresis: Used to determine protein concentration based on the differing mobility in an electric field.
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Urinary Protein (Albumin) Measurement: Measures protein (specifically albumin) in the urine. Normal urine does not have detectable proteins; high levels can indicate renal (kidney) disease.
- Methods: Urinary test strips to detect protein, the Biuret test, heat coagulation test, or more specific immunochemical /colorimetric methods.
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Urine Test Strips Method for Protein:
- Reagent: Tetra-bromophenol blue.
- Indicator: Urine strip turns green if protein is present.
- Sensitivity: Sensitive to albumin, less sensitive to hemoglobin and globulins.
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Heat coagulation test: This test involves heating the urine sample and causing proteins to precipitate (cloud or form a solid precipitate). If there is a cloudiness, it indicates protein.
Plasma Enzymes
- Functional Plasma Enzymes: Present in higher concentration in plasma compared to tissues; have known functions that depend on their role in the body, and remain present in the blood. Examples include clotting factors, lipoproteins, and pseudo-cholinesterase.
- Non-Functional Plasma Enzymes: Present in very low concentration in the plasma; unknown functions, often present in the blood because they are released from other tissues. Examples include ALT, AST, CK, LDH, alkaline phosphatase, acid phosphatase, amylase.
- Importance in Diagnosis of Disease: Levels of certain enzymes can indicate different diseases and pathologies that effect these organs. For instance, high levels of liver enzymes may suggest liver damage, inflammation, or disease. High levels of heart enzymes might indicate a heart attack or heart disease.
Enzyme Activity Factors
- Enzyme concentration: Increased concentration generally increases reaction rate.
- Substrate concentration: Increased substrate levels increase reaction rate until the enzyme is saturated
- Temperature: Enzymes work best within a suitable range of temperature
- pH: Enzymes have optimal pH range
- Co-enzymes: Some enzymes require co-enzymes (like vitamins, for reactions)
- Ion Activators: Some enzymes require ions to perform reactions.
- Inhibitors: Inhibitors delay or stop enzyme reactions
- Time: Reaction rates and duration can vary.
- Amylase: Enzyme responsible for converting starch to maltose; operates optimally at 37°C (body temperature).
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