Application Note

Washing MAGPIX beads with the AquaMax 2000/4000 Microplate Washer and LifeSep magnetic plate holder

  • Optimized washing for high-throughput bead-based multiplexing
  • Excellent retention of washed beads
  • Highly adjustable wash settings for maximum flexibility

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Introduction

The MAGPIX multiplexing system offers high-throughput multiplexing capabilities with up to 4800 results in one hour. To ensure optimal performance of these assays, it is essential to wash the MAGPIX beads efficiently while maximizing retention of the beads within the microplate wells. With the AquaMax® 2000/4000 Microplate Washers from Molecular Devices®, bead retention of at least 95% is achievable using the optimized settings described in this document.

magapix-beads-one

Figure 1. AquaMax 4000 Microplate Washer using the MAGPIX multiplexing system and a LifeSep magnetic plate holder.

Materials

Methods

Step 1. Diluted beads 1:50 in PBS and pipetted 100 μL beads into each well of a 96-well microplate.

Step 2. Read the microplate on the ImageXpress Velos Cytometer prior to washing to obtain the starting number of beads.

Step 3. AquaMax Washer microplate setup: Created a new plate in the onboard software. Started with 96-well Greiner plate, entered height as 24.2 mm (regular plate height + 10 mm LifeSep adapter height). Used the Well Depth Auto feature to determine the precise well depth and saved the resulting value in the plate definition.

Step 4. Washed plate with PBS on the AquaMax 4000 Washer. The following parameters gave the best results (minimal loss of beads during wash):

Aspirate

Dispense

With the ImageXpress Velos Scanning Cytometer, fluorescence was scanned using the 532 nm laser. The number of objects detected per well before and after washing was determined using the ImageXpress Velos Cytometer software.

Pre-wash bead counts for 96-well microplate

Average number of beads per well = 615

1
2
3
4
5
6
7
8
9
10
11
12
A
603
618
639
607
588
642
586
536
612
610
626
587
B
643
652
641
623
630
607
574
657
621
623
622
636
C
621
585
650
598
638
550
604
632
621
577
586
605
D
592
607
633
595
633
620
592
633
621
622
578
591
E
589
651
635
648
629
617
631
634
610
590
610
610
F
577
596
628
574
598
575
643
623
615
602
602
546
G
639
602
600
645
682
608
585
622
600
652
600
596
H
658
634
635
642
643
643
629
647
622
604
641
628

Post-Wash bead counts for 96-well microplate

Average number of beads per well = 585
% Retention = 95.1

1
2
3
4
5
6
7
8
9
10
11
12
A
584
572
599
597
580
621
554
552
608
605
566
515
B
634
630
643
548
622
579
608
613
535
540
598
616
C
563
590
641
591
602
575
560
594
622
533
576
554
D
495
593
568
513
605
600
587
551
573
577
499
568
E
527
618
607
634
578
593
605
604
640
575
479
539
F
575
540
621
576
595
554
607
605
617
603
574
586
G
544
577
561
567
630
621
625
595
594
626
571
506
H
608
606
546
616
625
599
628
625
611
566
599
613

magpix beads

Figure 2. Images of pre- and post-wash representative wells. A. Pre-wash representative well: d5, 633 objects (beads) counted. B. Post-wash representative well: D5, 605 objects (beads) counted.

Conclusion

A high rate of bead retention is essential for successful MAGPIX multiplexing assays. When MAGPIX beads were washed in a 96-well microplate on the AquaMax 4000 Microplate Washer, using the LifeSep magnetic plate holder and optimized settings as described above, 95% of beads were retained in the wells. The AquaMax Washers’ highly adjustable settings and interchangeable wash heads offer the ultimate in flexibility and optimization for any assay.

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