Applied Biosystems 7500 Getting Started

Chapter 3 Prepare the Reactions
Prepare the Standard Dilution Series
47
Applied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Standard Curve
Experiments
Notes
Prepare the Standard Dilution Series
Prepare the standard dilution series using the volumes that were calculated by the 7500
software (“Complete the Reaction Mix Calculations Tab (see page 33)” on page 32).
About the
Example
Experiment
For the standard curve example experiment:
• The standard concentration (stock) is 20,000 copies/µL.
• The volumes calculated in the software are:
Required
Materials
• Water (to dilute the standards)
• Microcentrifuge tubes
• Pipettors
• Pipette tips
• Standard stock
•Vortexer
• Centrifuge
Prepare the
Standard Dilution
Series for the
RNase P Assay
1. Label a separate microcentrifuge tube for each standard:
• RNase P Std. 1
• RNase P Std. 2
• RNase P Std. 3
• RNase P Std. 4
• RNase P Std. 5
Dilution
Point
Source
Source Volume
(µL)
Diluent Volume
(µL)
Total Volume
(µL)
Standard Concentration
(ng/µL)
1 (10,000) Stock 3.63 14.52 18.15 4000.0
2 (5,000) Dilution 1 9.08 9.08 18.15 2000.0
3 (2,500) Dilution 2 9.08 9.08 18.15 1000.0
4 (1,250) Dilution 3 9.08 9.08 18.15 500.0
5 (625) Dilution 4 9.08 9.08 18.15 250.0

Applied Biosystems 7500 Specifications

General

Detection Method	Fluorescence
Sample Capacity	96-well plate
Thermal Cycler	Yes
Temperature Uniformity	±0.5°C
Type	Real-Time PCR System
Thermal Accuracy	±0.25°C
Detection Channels	5
Weight	45 kg
Power Requirements	100-240 V, 50/60 Hz
Dynamic Range	9 logs
Reaction Volume Range	10-100 µL
Temperature Range	4°C to 100°C
Thermal Cycler Block	96-well
Software	SDS Software
Detection	Multiplex (up to 5 colors)