PRECISION OF DISPENSING
USING V&P WANDS AND MANIFOLDS
Paper by Duong Chau V&P Scientific Staff Scientist
Scientific wands and manifolds are designed for the purpose
of aspirating from microplates, washing microplates, and
dispensing into microplates. This experiment was set up to
determine the precision of the latter capabilities by
measuring the quantity of aqueous FITC dispensed per well
and calculating the coefficient of variations among the
wells as well as among the dispenses.
and manifolds range between 4 and 48 tubes and are designed
for microplates with wells between 48 and 1536. This assay
evaluated the performance of four separate devices, the
180, VP 184 (96 well format),
VP 189L, and
VP 189M (384 well
format); devices that represent the fundamental design of
the wands and manifolds. The experiment also provided the
data necessary to determine if the design differences
between wands and manifolds affect the dispense performance.
was done in a non-contact fashion (above the wells) and
generally resulted in coefficient of variations below 5% for
all devices tested. It was also observed that despite the
differences between the wands and manifolds in design, there
were no apparent differences in the quality of dispensing
capabilities. Only four devices out of many different
possible wand and manifold products V&P Scientific carries
were tested. However, this assay provides a proof of
principle for the design of the wands and manifolds, thus
similarly designed products are expected to yield similar
Scientific carries a variety of row/column wands and
manifolds designed for aspirating from and dispensing into
microplates. Manifolds are designed to have the
inlet/outlet port located at the top center of the barrel
while wands are designed to have to have the port located at
the end of the barrel. This allows a portion of the
wand to act as a handle for ease of handling the device.
Both styles, however, have anywhere between 4 and 48 tubes
designed for microplates with wells between 48 and 1536.
and manifolds can be connected to a vacuum line for
aspiration or used in conjunction with a liquid dispenser
such as a syringe, bottle top dispenser, or directly
connected to an elevated source. The focus of this
paper will be regarding the dispensing capabilities of the
wand and manifold using four different instruments
representative of the fundamental designs; the VP 180, VP
184, VP 189L, and VP 189M. The VP 180 is an 8-channel
manifold for 96 format microplates, the VP 189M is a
16-channel manifold for 384 format microplates, and the VP
184 and VP 189L are the wand versions respectively.
applications involving these devices generally do not
require exact precision, such as aspirating media from wells
or washing microplates. However, some applications
require that the instrumentation used to be precise and have
a low coefficient of variation. The precision of the
wands and manifolds was investigated as well as observing
any potential variation between the wands and manifolds for
liquid dispensing due to design differences.
Seen here is a VP 180 manifold.
Note that the port is positioned at the middle of the
Photo of VP 185 wand.
Note that the port is positioned at the end of the
of testing the wands and manifolds are fluorescence-based
readings, using fluorescein isothiocyanate (FITC).
FITC is a derivative of fluorescein with the molecule
functionalized with isothiocyanate, thus making it reactive
towards amine and sulfhydrl groups, and a great tool for
labeling proteins. FITC has an excitation and emission
wavelengths of approximately 495 nm/521 nm, and a maximum
energy output at pH. 8.0. Because FITC is pH sensitive, it
is important to keep it in a buffer to avoid even small
shifts in pH1.
excitation value output of FITC is directly proportional to
the amount of FITC molecules, hence a linear relationship
between the amount of FITC and the excitation signal.
Using this model, the experiment is designed such that the
amount of FITC dispensed by the manifolds in each well is
read, thus each well can be compared to one another.
The percent coefficient of variation (%CV) can be
extrapolated from this data using the formula:
coefficient of variation is a recognized value in the
laboratory for determining precision of instruments, thus
allowing for comparisons to other equipments with similar
functionalities (i.e. comparing the manifolds to
multi-channel pipettors commonly found in laboratories)2.
8-Channel Manifold (V&P Scientific VP 180)
8-Channel Wand (V&P Scientific VP 184)
16-Channel Manifold (VP Scientific VP 189M)
16-Channel Wand (V&P Scientific VP 189L)
Bottle Top Dispenser (V&P Scientific VP 195D-1)
96-well Polystyrene Black Assay Plate (Greiner Bio-One.
384-well Polystyrene Black Assay Plate (Greiner Bio-One
Microplate Reader (Victor 3)
Fluorescein 5-isothiocyanate, Isomer I (Sigma F7250)
Sulfoxide (Sigma D2650)
1 M Stock Solution, pH 8.0 (Sigma T3038)
milligrams of FITC was dissolved in 4 mL of DMSO and left
overnight to ensure complete dissolution and equilibration
of the mixture. Mix 80 uL of the FITC mixture with
2000 mL Tris-HCl, 0.1M, pH 8.0, buffer to get a final
concentration of 0.001 mg/mL FITC solution. This is
the solution to be dispensed during the assay.
concentration to be used is a function of several factors,
the most important to consider are the following; the
readable linear range of the plate reader, the strength of
the fluorescence, and the volume dispensed into the wells.
A standard curve was generated using a 12-channel pipette to
determine the ideal concentrations and volume of FITC to be
used, hence the concentrations used here reflects the
equipment and reagents used here.
and manifolds were set up according the manufacture’s
technote. The FITC solution was dispensed into 96-well
microplates using the VP 180 and VP 184 at volumes of 125 uL
and 250 uL per well. Each volume was tested in three
replicates (triplicate) resulting in a total of twelve
individual replicates. The VP 189M and VP 189L were
tested at a volume of 62.5 uL per well. Each device
was tested in replicates of twelve resulting in a total of
twenty-four individual replicates. All dispenses were
non-contact as the tubes neither touched the liquid nor
represents the data obtained using the VP 180 dispensing at
125 uL per well and 250 uL per well. The CVs ranged
between 3.19% and 9.75% for the 125 uL dispense, averaging
6.48%. The CV for one of the replicates was
particularly high due to one well (highlight) with
significantly lower readings. The average CV
disregarding the highlighted cell would be 3.38%. The
CVs ranged between 2.35% and 2.82% for the 250 uL dispense,
represents the data obtained using the VP 184 dispensing at
125 uL and 250 uL per well. The CVs ranged between
1.86% and 2.96% for the 125 uL dispense, averaging 2.46%.
The CVs ranged between 0.87% and 1.59% for the 250 uL
dispense, averaging 1.26%.