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Lab 1b_ImageJ Intro_TA Guide

Page history last edited by Kim Moore 7 years, 8 months ago

Phys 131, Fall 2012 TA GUIDE—Lab 1, Part 2

Lab 1(Part 2)—Introduction to ImageJ and Analysis of Cell Motion

Overview:

This is the second week of a two-week lab sequence designed to introduce the students to Excel (last week) and ImageJ. The students have completed an ‘Introduction to ImageJ’ reading for homework prior to this lab to familiarize themselves with the background and possible applications of ImageJ. This reading did not contain any technical knowledge about the operation of the program. To introduce them to the operation of ImageJ, they will be asked to analyze two of three cell motion videos and collaborate with another group to investigate a scenario (see below). The lab handout will give explicit instructions on the operation of ImageJ, but no guidance in the performance of the physical analysis. Your tasks are: 1) to demonstrate ImageJ skills when asked and 2) to act as a guide for the analysis (see the document on helping student groups learn). (It is often a good idea to put the approximate timing on the board, as well as a short list of the objectives/skill goals—see the following pages.) From a Biology perspective, we are hoping the students will: a) examine some of their underlying assumptions about the behavior of white blood cells and bacteria and b) root out misunderstandings fostered by cell videos that are often sped up or slowed down such that they seem as if they are occurring at the same rate.

Scenario:

A patient has a wound, in the process of healing, that is infected with bacteria. Will the patient need antibiotics?

Materials:

  • On the computer desktop:

    • ImageJ and appropriate plugins

    • Video files for qualitative analysis

      • Wound healing: WoundHealing.avi

      • White blood cells: Neutrophils.avi

      • Bacteria: E_Coli.avi

    • Video files for quantitative analysis (smaller chunks of the larger videos)

      • Wound healing: WoundHealing_25fps.avi, All students analyze this!

        • Technical specifications of video: 0.65 µm/pixel, 6.0 min/frame, playing at 25 frames/sec

      • White blood cells: Neutrophils_25fps.avi, Half of the groups analyze this.

        • Technical specifications of video: 1.326 µm/pixel, 7.2 sec/frame, playing at 25 frames/sec

      • Bacteria: E_Coli_25fps.avi, The other half of the groups analyze this.

        • Technical specifications of video: 156 pixels/µm, 0.050 sec/frame, playing at 25 frames/sec

  • 1 per person—ImageJ instructions/lab guide

Introduction:

  • Groups of four students—begin performing in the Community Lab roles for this lab, but make sure that each student gets a chance to operate ImageJ. (Groups of three are okay, but groups of five are usually too large.)

  • Explain the scenario. Discuss with students the basics of wound infection. What is the body’s immune system response? What might affect a decision to prescribe/not prescribe antibiotics? Show the long videos (or have the students view them on their desktops). What do the videos show us? How does this affect the antibiotics decision? Motivate the need for quantitative analysis of the videos.

  • Explain that everyone will analyze the 1st video (wound healing) and then they will split the remaining two videos—half the class will look at white blood cells and the other half will look at bacteria. They will then confer with a group that studied the other video and come to a decision about the antibiotics. Once the groups have decided, a class discussion will take place.

  • Briefly review the Skill Goals (Biology and ImageJ, on the next page) and remind the students that they are to recall and practice the Physics and Excel skills learned last week.

  • Remind students that they are ALL expected to master the skills, so they should take turns and help each other out. Taking notes may not be a bad idea, either.

  • Inform students that a lab report (discussing their qualitative and quantitative exploration of and resolution to the scenario) will be collected from each group at the end of the lab today. They may find it helpful to write portions of the report as they go, documenting their evolving understanding of the problem investigation.

Interim Discussion:(after the wound healing video & before they look at the bacteria/neutrophils)

  • What difficulties/choices did the students encounter in analyzing the wound healing video?

  • Explain once more that they will split the remaining two videos—half the class will look at white blood cells and the other half will look at bacteria. Assign the groups to the videos. They will then confer with a group that studied the other video (assign the companion group pairings) and come to a decision about the antibiotics. Once the groups have decided, a class discussion will take place.

Summation/Submission:

  • Have the student groups briefly share and explain their decision/resolution to the scenario. If different groups come to different conclusions, discuss why this has happened. What other factors could affect the need for antibiotics? (Having the class come to the conclusion that more quantitative information is needed before the decision can be made is a perfectly acceptable result. The goal is to get them thinking critically about the question and their own assumptions.)

  • Discuss the challenges and considerations with the class, if any have not yet been addressed. Ask the students what they found most difficult/challenging about either the ImageJ operation or the cell motion analysis.

  • With approximately 20 minutes remaining, direct the students to finalize their group report (discussing their qualitative and quantitative exploration of and resolution to the scenario). (Each sub group in the companion group should write their own report—so one report for each group of four students.) They can reference the Scientific Community Lab document if they need more information about the structure and purpose of the report.

  • At the end of the lab, collect the reports from each group (make sure that the group members have written all their names on the front page). These should be graded by you according to the Community Lab rubric. Good feedback now will save you time later.

Biology Skill Goals:

Challenges/Considerations:

Qualitatively compare videos of motion for cell sheet migration, neutrophils, and bacteria

Do they look like they are happening at the same rate? What else do you know about these processes that might help you make a prediction?

Understand the need for quantitative analysis

What are the possible results given different speed rankings? (E.g., if the white blood cells move fastest of all while actively seeking the bacteria, then antibiotics are not likely to be necessary, regardless of the speed of wound closure. If all speeds are about equal, can we answer the question or will we need more knowledge? If the bacteria are the fastest, will the wound closure rate matter?)

Quantitatively compare these videos of motion

Are you surprised? Is this what you expected? How does this change your understanding of the qualitative aspects of the videos?

Use the quantitative analysis to make a prediction and determine what other factors might need to be investigated

Will antibiotics be necessary? Why?/Why not?

What else could affect the need for antibiotics?

(Health of patient (immune-compromised? maturity and density of white blood cells), type and resistance of bacteria, concentration of bacteria, toxicity of phagocytized bacteria) Do bacteria actively evade the white blood cells?

 

 

Recall and Practicethe Physics Skill Goals & Excel Skill Goals from Lab 1

ImageJ Skill Goals:

  • Open a video file in ImageJ

  • Understand the vocabulary of ‘frame’ and ‘stack’ and ‘image sequence’

  • Choose which objects to track (How many do we need to track? What part of the object are we tracking? How do we pick a good candidate for tracking? What makes a candidate good/bad? What challenges exist for tracking the object?)

  • Operate the Manual Tracking plugin (What physical steps are required to collect data? Where is that data? How do we review our work? Tracking multiple objects. Tracking objects with different start/stop times.)

  • Understand the ‘centring’ option (What is its function? What are its limitations? Adjusting ‘search square size for centring.’)

  • Export the data to Excel

  • Understand the challenges of interpreting the data within Excel (How do we determine the correct time for a data point? Can we use the velocities (actually speeds) given by Manual Tracking? How do we convert the x and y pixel coordinates into positions? Do we need to consider both the x and y motions? Why?/Why not?)

Approximate Timing:(2 hours)

  • Introduction: .…………………………………………………… 10 minutes

  • Data Collection & Analysis, 1st video: ……………………………. 30 minutes

  • Class Discussion of 1st video: ……………………………………. 5 minutes

  • Data Collection & Analysis, 2nd video: ……………………………. 30 minutes

  • Conferring with companion group, comparing all three videos: 10 minutes

  • Class Discussion/Summation: ……………………………………. 10 minutes

  • Finalizing Lab Report: ……………………………………………. 25 minutes

 

 

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