Description

Single Cell Viability Analysis by Impedance Flow Cytometry - Ampha Z32

 

Amphasys’ new-generation flow cytometer is a plug & play device designed to blend simplicity with flexibility. The absence of optical components minimizes set-up times and maintenance to a minimum. Time-consuming sample preparation procedures and expensive fluorescent dyes do not belong to Amphasys’ world, because it is exclusively founded on electrical measurements. Latest generation digital signal processing electronics are specifically tailored to Amphasys’ semi-disposable chips, such that best-of-class impedance analysis on single cells is achieved.
The small dimensions of Ampha Z32 and the robustness of the electrical measurement provide a mobile and portable solution for any kind of cell analysis, be it in your laboratory or out-side in the field. Designed initially for research purposes, Ampha Z32 is now particularly suitable for routine applications, where detailed, high-quality cell analyses and ease-of-use are major requirements.

 

Chips

The heart of the cytometer consists of a microfluidic chip equipped with microelectrodes. This high-sensitivity sensor measures changes of the electrical resistance of the fluidic medium when cells or other particles pass through the applied AC electric field.

In order to ensure highest possible sensitivity Amphasys has developed a number of chips with different channel dimensions ranging from 15 to 250 μm. The recent production of the new 250 μm chip marks a milestone in chip design and is a breakthrough for the application to large pollen. The whole range from very small cells like bacteria to very large cells such as certain pollen grains like pumpkin, maize or cotton can now be analyzed. Chips can be used for up to several hundred measurements when treated with care.

 

Reagents and Disposables

Amphasys provides specialized reagents and consumables for many kinds of cells and applications as well as cleaning solutions for the care of the fluidic system.

The electric signal recorded by the highly sensitive electrodes is influenced by the cell itself and by the surrounding liquid. The contents of the buffers may have an effect to the biochemical behavior of the cells as well as the conductivity of the liquid and therefore on the signal. For these reasons, the conditions of the buffers including solvents, additives, salts, pH etc. have been optimized to keep cells viable and to obtain highest measurement sensitivity at the same time.

 

Mobility Package

Ampha Z32 is designed to bridge the gap between the lab and the field. Its weight of about 8 kg and dimensions of 25 x 27 x 35 cm make it perfectly suited as a table-top lab instrument as well as a mobile field instrument. With the optionally available mobility package including a 24 V rechargeable battery and a 24 V truck adapter cable it can be used independent from a power outlet.
The instrument is of a robust build and can easily be transported in the trunk or on the back seat of a car. However, we also provide a heavy duty aluminum transport casing if you plan to move the instrument on a regular basis.

 

Tutorial Videos

To view tutorial videoes press here>>

 

Applications

Breeding

Temperature, humidity and factors like the application of pesticides or inbreeding depression have a direct impact on pollen viability, quantitiy and seed yield. 

Now, with the Ampha Z32, you can get a precise measure of the pollen viability along your pollen supply chain in the lab or greenhouse, while eliminating time-consuming and inaccurate germination and staining assays. 

Increase breeding efficiency and get to market earlier through: 

• Improving crossing success rates
• Maximizing the window for master lines
• Breeding for heat tolerance by selecting for pollen viability under heat stress
• Monitoring the sterility of cytoplasmic male sterile (CMS) lines

Hybrid Seed Production

Control over pollen quality and pollination along the pollen supply chain is a key success factor to ensure high quality seed and yield. 

Excessive warm nights, draughts, rain, and pesticide applications negatively affect the pollen viability, quantity, and timing of pollen shed, leading to a reduced seed set and yield loss. Consequently, measuring and understanding pollen quality in hybrid seed production allows maximization of yield, improvement of the female to male ratio and forecasting of your yield. 

Fruit and Nut Production 

Effective pollination is indispensable to ensure high seed and fruit yield. Asynchronous flowering of pollinator trees in the orchard, rising temperatures and draughts, as well as the decline of natural insect pollinators severely affect this biological process and cause significant losses in tree nut and fruit production. 

Research

Challenges in Pollen Analysis

 Traditional methods to determine pollen viability often rely on tedious sample preparation steps and labor intensive microscopy analysis. 

With the Ampha Z32, sample preparation is just a simple resuspension and filtration that takes less than a minute and works for all species (currently over 200 species tested). 

 

 

Pollen Viability & Seed Set

 

Seed set in tomato as a result of pollination with pollen of varying quality. 

Pollen viability measured on the Ampha Z32 before pollination: A = 31%, B = 3.5%, and C = 0.3%. The size of fruits resulting from pollinations with these pollen samples (pictured), as well as the seed set (graph) correlate with the pollen viability. 

Other species show similar behavior, althogh each species has individual correlation patterns. 

 

 

 

Pollen Viability & Germination

 

Pollen viability analysis with IFC and in vitro germination show an excellent correlation ( R² = 0.96). While over 10,000 pollen per minute were analyzed with the Ampha Z32, only 100 pollen could be counted in the same time with the germination assay. This high count results in a standard deviation of <1% for IFC

 

 

 

 

Pollen Count

 

Counting pollena as part of a pollen dispersion, pollen potential or pollen emission analysis is the standard method for calculating the number of males needed for pollination. 

An accepted method relies on collecting the pollen in Isoton media for preservation and counting with a Coulter Counter. The comparison between the Ampha Z32 and the Coulter Counter shows that besides providing viability data, the Ampha Z32 can at the same time also be used to rapidly count large amounts of pollen. 

 

 

Pollen Ploidy Analysis

 

 

Analysis of ploidy in Watermelon Pollen (Citrullus lanatus).

Diploid and tetraploid samples can be distinguished by analyzing their average size. Similar results were obtained for many other species like kiwi, tomatoes, potatoes, wheat, and cyclamen. 

 

 

 

 

Developmental Stages

 

 

 

No time-consuming DAPI staining is required to determine the stage in microscope development anymore! IFC on the Ampha Z32 is so sensitive, it allows to distinguish between unpolarized and polarized microspores, as well as bi-cellular and tri-cellular pollen. 

Besides staging, the Ampha Z32 allows early detection of induction success, allowing earlier selection of cultures. Optimize your DH induction protocols and monitor the induction success. 

 

 

 

Pollen Expert Blog

Overcoming the Pollination Gamble with Science - November 19, 2020

Measuring Wheat Pollen Viability - A Delicate Procedure?! - October 20, 2020

How Pollen Quality Affects Seed Set - September 22, 2020

Unravelling Hidden Secrets Behind Dead Tomato Pollen - June 4, 2020

Impact of Pollen Quality on Tomato Seed Production - February 28, 2020

 

Technical data

Impedance Chip
Available sensing channel dimensions	15 x 15 μm, 30 x 30 μm, 50 x 50 μm, 80 x 80 um,  120 x 120 μm or 250 x 250 μm (depending on application requirements)
Impedance measurement
Frequency range	0.1 MHz – 30 MHz
Frequency selection	Up to 4 different frequencies simultaneously
Analysis range
Sample volume	50-2000 μl (typically)
Concentration range	1 x 10(3) to 1 x 10(7)cells/ml
Particle size	1 - 60 μm (up to 150 μm with 250 μm chip)
Fluidics
Sample flow rate	5 – 2500 μl/min (depending on channel dimension)
Pump	Peristaltic pump with disposable pump head
Labware compatibility	Standard 5 ml polystyrene round -  bottom tubes (Falcon® 352058)
Operating System (min requirement)
	Laptop with Windows® 7 or 10
	15” TFT LCD display
	i5 or i7 QuadCore processor
	128 GB SSD
Dimensions & Weight
Dimensions (W x D x H)	255 x 275 x 353 mm
Weight	8.4 kg
Operating Environment
Temperature	16° - 32°C
Humidity	10% - 90% relative non - condensing
Power	24V DC ± 10% , max. 3A, < 90 W