An image needs to be made transparent so that it will appear properly on a website. Also, it needs to have thousands of different colors in it. Which file format would be best in this situation?

The file format that would be best in this situation is PNG (Portable Network Graphics).

PNG supports transparency and is commonly used for web graphics and images that require transparency effects. With PNG, you can make the background of an image transparent, allowing it to blend seamlessly into any website background.

Furthermore, PNG format supports millions of colors and can store images with high color depth, making it suitable for an image with thousands of different colors.