Laboratory Training

Summer Internship 2015 ACRM

Pipetting

• Volumes: 1000µL, 100µL, 10µL • 15 repeats each • Acceptable variation 5%

Method • Pipette distilled H20 onto a balance and

record the weight

Microbeads

• Concentrations: High, Low • 3 repeats each • Acceptable variation 15%

Method • Fill microcell and perform count in 4 fields • Calculate average and concentration • Validate against reference values

Sperm Concentration

• Evaluate 10 Semen samples • Acceptable variation 20%

Method • Fill microcell and perform count in 4 fields • Calculate average and concentration • Validate and compare to mentor’s results

Sperm Motility

• Evaluate 10 Semen samples • Acceptable variation 20%

Method • Fill microcell and perform count in 4 fields • Calculate average and % motility • Validate and compare to mentor’s results

Sperm Vitality

• Prepare 2 different smears • Evaluate 5 reference slides • Acceptable variation 15%

Method • Evaluate each slide x 2 • Count viable and non-viable cells (100 sperm) • Calculate average and concentration • Validate and compare to reference values

Data Recording

• Record all information in your lab book

• Transfer to day sheet

• Populate soft copy (Excel)

• Print and have it verified

Summer Internship Bench Research Training

The interns will observe and practice the following techniques prior to the actual start of their original research projects.

Laboratory safety:

• Explain the dress code that needs to be observed (lab coats, closed shoes, etc.), designated work areas, and the basic safety procedures to be followed.

Pipetting:

• Explain the different types of pipettes (transfer pipette, micropipette, pipettor), when they should be used and how they should be operated.

• Exercise - Pipette 3 volumes (1000µL, 100µL and 10µL) x 15 repeats

Microscopy:

• Explain the various types of microscopes, the function of different components as well as the proper operating and cleaning procedures (Tutorial)

General handling of a semen sample:

• Explain and demonstrate the basics to be performed when a semen sample was received: Incubation, Recording of information, Liquifaction, transfer, volume, pH, Viscosity, Color

Microbeads:

• Explain the loading of a microcell, setting up of the microscope (x20 phase contrast), counting technique and calculations

• Exercise – Count 2 concentrations x 3 each

Sperm count and Concentration:

• Explain the loading of a microcell, setting up of the microscope (x20 phase contrast), counting technique and calculations

• Exercise – Analyse 10 x samples

Vitality

• Explain the principles of employing a dye exclusion staining technique for viability measurement, preparing the sample, making of the smear, coverslip slide, and evaluating the sample

• Exercise – Prepare and score x 5 slides

Morphology:

• Explain the principles of morphology, the preparation, staining and evaluating of slides

• Demonstrate – Slide preparation, staining, scoring

• Exercise – Only identified students will receive further training

TUNEL

• Explain the principles of TUNEL as well as the calculations and preparation method.

• Exercise – Only identified student will be receive further training

ROS

• Explain the principles of ROS measurement

• Demonstrate – Perform the sample preparation, machine setup and calculation

• Exercise – Only identified student will be receive further training

ORP

• Explain the principle of sORP and cORP

• Demonstrate – Perform sample preparation, machine setup and calculation

Specialized Techniques

• Techniques specific to projects (e.g. Acrosin activity, Western Blot analysis, Proteomics, HOS, etc.) will be explained by the particular mentors to the individual(s) responsible in each group

Summer Internship 2015

Bench Research Training Proficiency Report

Intern: Mentor:

Pipetting Proficiency Pipette: P-1000 Reading Actual

volume (µL)

Measured volume

(µg)

Absolute Difference

Percentage Difference

(%)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Average Coefficient of Variance (CV)

Acceptable variation: 5% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)

Pipette: P-200 Reading Actual

volume (µL)

Measured volume

(µg)

Absolute Difference

Percentage Difference

(%)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Average Coefficient of Variance (CV)

Acceptable variation: 5% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)

Pipette: P-200 Reading Actual

volume (µL)

Measured volume

(µg)

Absolute Difference

Percentage Difference

(%)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Average Coefficient of Variance (CV)

Acceptable variation: 5% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)

Counting Microbeads: Low concentration Reading Reference

Count (x106/ml)

Measured Count

(x106/ml)

Absolute Difference

Percentage Difference

(%)

1

2

3

Average

Coefficient of Variance (CV)

High concentration Reading Reference

Count (x106/ml)

Measured Count

(x106/ml)

Absolute Difference

Percentage Difference

(%)

1

2

3

Average

Coefficient of Variance (CV)

Acceptable variation: 15% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)

Sperm concentration: Reading Reference

Count (x106/ml)

Measured Count

(x106/ml)

Absolute Difference

Percentage Difference

(%)

1 2 3 4 5 6 7 8 9 10 Average Coefficient of Variance (CV)

Acceptable variation: 20% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)

Motility: Reading Reference

Motility (%)

Measured Motility

(%)

Absolute Difference

Percentage Difference

(%)

1 2 3 4 5 6 7 8 9 10 Average Coefficient of Variance (CV)

Acceptable variation: 20% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)

Vitality (E&N): Reading Reference

Viability (%)

Measured Viability

(%)

Absolute Difference

Percentage Difference

(%)

1

2

3

4

5

Average

Coefficient of Variance (CV)

Acceptable variation: 15% Met the criteria: Yes / No Retraining completed: Not Applicable / Yes / No Verified: Approved: (Mentor) (Director)