Blood Lab - Differential White Blood Cell Count

Key Concepts

• Leukocytes, or white blood cells (WBCs), can be divided into two subgroups depending on whether or not their cytoplasm contains granules that are visible using a brightfield microscope.

  • Agranulocytes do not have visible granules.

    • Lymphocytes: Small cells with a round nucleus taking up most of the volume of the cell.

    • Monocytes: Large cells with a bean-shaped nucleus taking up half to nearly all the volume of the cell.

  • Granulocytes contain granules that stain characteristic colors.

    • Basophils: Contain dark purple or black granules. The nucleus is usually bilobed (two lobes).

    • Eosinophils: Contain bright red granules. The nucleus is usually bilobed (two lobes).

    • Neutrophils:Contain light pink and/or purple granules. The nucleus is multilobed (multiple lobes).

• A differential white blood cell count (DIFF) determines the percentage of the various types of white blood cells (WBCs), also known as leukocytes, in a blood sample.

• A complete blood count (CBC) with DIFF is often run in the lab on a machine that counts with an algorithm to match cells. However, many clinicians request a manual count from a trained lab tech's perspective when highly accurate counts are necessary or if cells are expected to be irregular, such as in leukemias.

• It is important to learn this technique, as the values that come back from this test are important to clinical diagnosis and evaluation of how/what the immune system is doing in the patient.

• Normal DIFF counts are approximately as follows:

  • Neutrophils 40-70%

  • Lymphocytes 20-40%

  • Monocytes 2-8%

  • Eosinophils 1-4%

• Reference ranges can vary by age, sex, methods of testing, and other factors. There are no nationally established reference ranges for DIFF values; instead, each laboratory tests a population and establishes its own reference ranges. Therefore, the reference ranges quoted are only approximate.

• Reference ranges can vary by age, sex, methods of testing, and other factors. There are no nationally established reference ranges for DIFF values; instead, each laboratory tests a population and establishes its own reference ranges. Therefore, the reference ranges quoted are only approximate.

Overview

• In this simulation, you complete a differential white blood cell count (DIFF) manually using a prepared blood slide and a brightfield microscope. It is helpful to understand the manner in which this process works and to further familiarize yourself with WBC appearance.

• To prepare for the count, you will first identify the 5 types of WBCs and the lab equipment used to perform the DIFF.

• Next, you will prepare your own wet mount of blood using the Wright stain technique.

• A DIFF can only be performed using high magnification so the types of WBCs can be distinguished.

• Take a moment to reflect on personal safety precautions. Working with blood is a potentially hazardous situation. In real life, you should:

  • Wash the laboratory lab benches before and after the procedures with an appropriate disinfectant.

  • Wear disposable gloves, lab coat, and goggles when handling blood samples.

  • Wash your hands after the laboratory.

  • Only use a blood lancet once.

  • Dispose of used lancets, pipettes, and other blood contaminated items in the appropriate hazardous waste container, never a regular trash container.

Before you begin

• Knowledge of the operation of a brightfield microscope is assumed for this simulation.

• A good blood smear is necessary for accurate counts.

• You will slowly move the slide top to bottom to identify and count 50 WBCs, then calculate the percentage of each type of WBC.

• Only count the middle of the blood smear where the red blood cells just nearly touch each other and there is little white space.

• Near the edges of the smear, the cells will either overlap or be spread out too much for an accurate count.

  

• How to move the slide: Make sure to only count each area once by moving the slide in a straight line through one field of view at at time.

Laboratory Simulation

Methods

• Phase 1: WBC identification

  • Identify different WBCs (leukocytes)

• Phase 2: Lab equipment

  • Identify lab equipment

• Phase 3: Preparation of blood smear

  • Place microscope slide on staining rack

  • Add drop of blood to slide using pipette. Dispose of pipette

  • Take another slide from box and use it to smear blood drop by sliding it from right to left. Let air dry for 1 minute. Dispose of slide

  • Add Wright stain to smear. Wait 2 minutes

  • Add distilled water to smear. Wait 4 minutes

  • Wash slide with filtered water. Wait 1 minute

  • Place slide on microscope stage

• Phase 4: Microscope differential count

  • Focus slide at 4X using the coarse focus, then the fine focus

  • Turn objective to 10X and focus again

  • Move to upper-left corner of slide

  • Open Lab Data. Count each type of WBC in each region of slide by clicking corresponding buttons in Lab Data.

  • Count 50 WBCs in total by using arrow buttons to move through regions of slide.

• Phase 5: Lab wrap-up

  • Select your answer to the question

• Phase 6: Save Lab Data

  • Relevant Lab Data is available to be saved for personal reference. Data will be available if you return to this laboratory simulation

White Blood Cells

Supplies

Collected Data