BlockIt™ Plus microarray blocking solution designed to inactivate reactive groups remaining post-printing on SuperEpoxy, SuperAmine, SuperAldehyde, SuperNitro, SuperPVDF and SuperNylon and other microarray substrate slide surfaces. BlockIt™ Plus Blocking Solution greatly reduces background noise while maintaining full signal intensities for a wide spectrum of microarray applications. 250 ml of 1X buffer.

Table of Contents

• Introduction
• Quality Control
• Product Description
• Technical Assistance
• Short Protocol
• Complete Protocol
• Short Protocol DNA
• Complete Protocol DNA
• Troubleshooting Tips
• Recommended Requirements
• Ordering Information
• Warranty

Introduction
Congratulations on taking a big step towards improving the economies of scale, quality and speed of your microarray research. This booklet contains a complete set of protocols outlining the steps and principles needed to use BlockIt™ Plus blocking buffer for microarray surface chemistries. This product is design for use prior to incubation and hybridization reactions involving protein and DNA microarrays.

Quality Control
Arrayit assures the performance of these products, the finest scientific research went into their development. Store at 4°C, the product has a 1-year shelf life when handled and stored properly.

Product Description
Arrayit BlockIt™ Plus blocking buffer is designed for blocking the portions of microarrays that are still reactive after printing to eliminate fluorescent background following the incubation or hybridization reaction of protein and DNA microarrays. Users will appreciate the following features:

• Often works when regular BlockIt™ is not sufficient
• Ultra-pure reagents used for buffer formulation
• Provided as 1X ready to use concentrate
• Recommended for use on all types of protein or DNA microarray glass microarray surface chemistries
• Easy cover slip or petri dish blocking procedure for minimal reagent consumption
• 250 ml volume provided can efficiently block thousands of microarrays
• Will not cross react with proteins and DNA

Technical Assistance
Please contact us if you have any comments, suggestions, or if you need technical assistance. By electronic mail: arrayit@arrayit.com (under the subject heading, please type Arrayit technical assistance). By telephone: (408) 744-1331, Monday–Friday PST 9:00am - 4:30pm. Please remember that we want to hear about your successes!

Short Protocol for Protein Microarrays
Wash microarray and then incubate microarray in 1X BlockIt™ Plus buffer under a coverslip for 1 hour prior to protein binding or hybridization reactions.

Complete Protocol for Protein Microarrays
BlockIt will couple to un-reacted binding groups on microarray surfaces and prevent background fluorescence. Once the printing process is complete, and samples are coupled to the surface, block a microarray surface by using a 1-24 hour incubation at room temperature to 4 degress celcius in 1X BlockIt™ Plus buffer. Perform this reaction under a coverslip using 4 ul of BlockIt for every cm2 of coverslip area, also compatible with LifterSlips. A Hybridization Cassette can be used to keep the blocking reaction from drying out. After blocking, wash the microarrays to remove excess buffer. Wash 2 times for 15 minutes each in a High Throughput Wash Station in Protein Microarray Wash Buffer to remove unbound protein molecules and buffer components from the microarray. Protein and DNA binding to the SuperEpoxy, SuperNitro and SuperPVDF surfaces is extremely stable and the microarrays can be washed, blocked and reacted without sufficient loss of coupled protein.

NOTE: BlockIt™ Plus is not recommended to be used as a reaction buffer for protein microarray applications.

Short Protocol for DNA Microarrays
Incubate microarray in 1X BlockIt™ buffer under a coverslip for 1-24 hours prior to hybridization.

Complete Protocol for DNA Microarrays
BlockIt will couple to the un-reacted groups on the surfaces and prevent background fluorescence. Once the printing process is complete, and samples are bound to the surface use a 1-24 hour incubation at room temperature of 1X BlockIt Plus buffer. Perform this reaction under a cover slip using 4 ul of BlockIt for every cm2 of coverslip area. A Hybridization Cassette can be used to keep the blocking reaction from drying out. After blocking wash the printed microarrays in a High Throughput Wash Station using the protocol recommended by the manufacturer. For best results use Arrayit DNA Microarray Wash Buffers. DNA binding to the SuperAmine, SuperAldehyde and SuperEpoxy surfaces is extremely stable and the microarrays can be washed, blocked and reacted without significant loss of coupled DNA. After blocking, wash and process the microarrays according to the manufacturers recommendation to remove excess buffer and unbound sample.

Figure 1. Shown is a microarray blocking reaction using BlockIt™ Plus Microarray Blocking Buffer.  Three DNA microarrays can be incubated overnight with gentle agitation in a square culture dish containing 30 ml of 1X BlockIt™ to inactivate the microarray surface and reduce fluorescent background during the hybridization reaction. BlockIt™ Plus molecules bind covalently to un-reacted groups on the microarray surface, preventing non-specific binding of probe molecules during hybridization. BlockIt™ Plus is not a clear solution, but contains highly purified and soluble components that provide a highly efficient blocking function.  BlockIt™ Plus is an all-purpose blocking solution for DNA, protein, and other types of microarrays and is used when blocking with BlockIt™ shows to be inefficient.

Troubleshooting Tips
If microarray shows high background:

• Confirm that the intrinsic background of the microarray is low to start with.
• Be sure to not let any sample dry on the microarray surface during the binding reaction.
• Make sure to purify away unincorporated dyes and other conjugates such as biotin, AP and HRP in the labeling reaction.
• Take care to block for the recommended time (at least 1 hour).
• Pay attention to the proper handling and shelf life of the buffer
  (1 year shelf life, Store at 4 C).
• Do not skip washing the microarray after the blocking step.

Recommended Equipment:

• Hybridization Cassette or Hybridization Cassette Plus
• High Throughput Wash Station
• Microarray Centrifuge